Quantification Of Neuronal Morphology And Maturation Of N2a Cells Using NeurphologyJ Software

Anthonysamy, Doreen Maria (2020) Quantification Of Neuronal Morphology And Maturation Of N2a Cells Using NeurphologyJ Software. [Project Paper] (Submitted)

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Abstract

N2a is a cell line used widely in studies of neuronal maturation and development in mouse neuroblastoma. Growth of N2a cells via its differentiation into mature neurons is traditionally induced by serum deprivation (SD) method as serum has been known to contain inhibitory factors that may impede neuronal differentiation and neurite growth. Even though that soma count and neurite length, amongst other neuronal features of differentiated N2a cells, can be efficiently-analyzed using various open-source tools and software, the use of NeurphologyJ software to characterise and examine the morphological features of N2a cell has not been explored within the neuroinflammation group. To measure neuronal morphology parameters (soma count, neurite length, neurite count, attachment point and endpoints) of undifferentiated and differentiated N2a cells using NeurphologyJ software. N2a cells were maintained in 0.5% fetal bovine serum at 37°C in DMEM supplemented and 5% CO2 incubation conditions and were stained with FITC-labelled neuron-specific class III beta-tubulin (Tuj1) marker and 4′,6-Diamidino-2-Phenylindole (DAPI), prior viewing under a fluorescent microscope. The cell culture was conducted by another colleague. Captured images were processed using a fluorescent imaging system, converted to TIFF file format, and were digitally-enhanced before evaluation using NeurphologyJ software. Optimal threshold levels were set individually for each parameter, prior features extraction and determination. These features include neurite length, attachment point, neurite count, endpoints, and soma count. The differences between undifferentiated and differentiated N2a cells were analyzed using the Mann–Whitney test. It was determined that differentiated cells when compared with undifferentiated cells, represent significantly (p<0.05) higher counts of soma (161.33±64.96 vs 88.50±23.81), neurite count (2362±605.43 vs 772.00±248.01), neurite length (2348.67±1345.79 vs 18451.33±7963.13), attachment points (995.67±295.88 vs 185.83±90.95), and endpoints (1383±622.28 vs 166.33±85.77). These findings suggest that NeurphologyJ software can be useful for the measurement of morphological features of both undifferentiated and differentiated N2a cells. For future applications, N2a cells differentiated using other types of methods may be conducted for a better understanding of cell maturation parameters. Other than that, a comparison of N2a cells with other neuronal cell lines by measuring the parameters using NeurphologyJ software can also be conducted.

Item Type:	Project Paper
Faculty:	Faculty of Medicine and Health Science
Depositing User:	Ms. Nor Safa'aton Saidin
Date Deposited:	23 Aug 2023 00:11
Last Modified:	23 Aug 2023 00:11
URI:	http://psaspb.upm.edu.my/id/eprint/1333

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