Measuring the Apoptotic Response of the HT22 Mouse Hippocampal Cell Line to Neuronal Differentiation Medium

Ayla Batu Öztürk; Derya Yetkin; Nail Can Öztürk

Article Details

Measuring the Apoptotic Response of the HT22 Mouse Hippocampal Cell Line to Neuronal Differentiation Medium

Authors:

Ayla Batu Öztürk | Derya Yetkin | Nail Can Öztürk

Publisher:

Sakarya University

Main Topics:

Sağlık Kurumları Yönetimi

Journal:

Sakarya Tıp Dergisi Vol.13 Issue 3, pp.375-383

Document Type:

Article

Publication Date:

9/2023

Document Language:

Turkish

Document ID (Org No.):

2001298974

pISSN:

2146-409X

Keywords:

Abstract:

 Objective: The aim of this study is to test the apoptotic response of the HT22 mouse hippocampal cell line, which is frequently used in neurobiology studies, to a standard differentiation medium at different times in order to understand its usability as a neuronal differentiation model. Methods: For seeding and culturing HT22 cells aside the differentiation, HG-DMEM and for differentiation B27 supplemented NB medium were used respectively. A total of 9 different groups, including the control group, were cultured in standard cell culture environment. At the end of the experiment, the rates of earlylate apoptosis and necrosis were determined with flow cytometric analysis of AnnexinVPI labeled HT22 cells. After testing whether the raw data fit the normal distribution or not by employing Shapiro-Wilks test, further statistical analyzes were performed with the most appropriate test parametric or non-parametric tests. Results: Statistically significant differences were found in terms of early apoptosis P0,01 and late apoptosis P0,001 cell population rates, while there was no significant difference of necrosis P0,064. Significant P0,001 increase in early apoptosis was evident in the majority of groups cultured differentiation medium for 48 or 72 hours, compared to the cells cultured with only HG-DMEM or differentiation media for 24 hours. While a significant increase in late apoptosis was observed in the majority of the groups cultured with the differentiation medium compared to the control group P0.001, no significant difference was observed between the groups differentiated for different durations P0,05. Conclusion: HT22 hippocampal cell line, which is frequently used in in-vitro modeling of important pathobiological features of neurodegenerative diseases, but poorly understood for the suitability in neuronal differentiation, has a markedly increased response to a widely used differentiation medium in terms of early apoptosis culturing duration dependent and late apoptosis regardless of duration. Our findings provide important clues at the level of apoptosis in terms of the usability of the HT22 cell line in differentiation studies.

 Objective: The aim of this study is to test the apoptotic response of the HT22 mouse hippocampal cell line, which is frequently used in neurobiology studies, to a standard differentiation medium at different times in order to understand its usability as a neuronal differentiation model. Methods: For seeding and culturing HT22 cells aside the differentiation, HG-DMEM and for differentiation B27 supplemented NB medium were used respectively. A total of 9 different groups, including the control group, were cultured in standard cell culture environment. At the end of the experiment, the rates of earlylate apoptosis and necrosis were determined with flow cytometric analysis of AnnexinVPI labeled HT22 cells. After testing whether the raw data fit the normal distribution or not by employing Shapiro-Wilks test, further statistical analyzes were performed with the most appropriate test parametric or non-parametric tests. Results: Statistically significant differences were found in terms of early apoptosis P0,01 and late apoptosis P0,001 cell population rates, while there was no significant difference of necrosis P0,064. Significant P0,001 increase in early apoptosis was evident in the majority of groups cultured differentiation medium for 48 or 72 hours, compared to the cells cultured with only HG-DMEM or differentiation media for 24 hours. While a significant increase in late apoptosis was observed in the majority of the groups cultured with the differentiation medium compared to the control group P0.001, no significant difference was observed between the groups differentiated for different durations P0,05. Conclusion: HT22 hippocampal cell line, which is frequently used in in-vitro modeling of important pathobiological features of neurodegenerative diseases, but poorly understood for the suitability in neuronal differentiation, has a markedly increased response to a widely used differentiation medium in terms of early apoptosis culturing duration dependent and late apoptosis regardless of duration. Our findings provide important clues at the level of apoptosis in terms of the usability of the HT22 cell line in differentiation studies.

 Objective: The aim of this study is to test the apoptotic response of the HT22 mouse hippocampal cell line, which is frequently used in neurobiology studies, to a standard differentiation medium at different times in order to understand its usability as a neuronal differentiation model. Methods: For seeding and culturing HT22 cells aside the differentiation, HG-DMEM and for differentiation B27 supplemented NB medium were used respectively. A total of 9 different groups, including the control group, were cultured in standard cell culture environment. At the end of the experiment, the rates of earlylate apoptosis and necrosis were determined with flow cytometric analysis of AnnexinVPI labeled HT22 cells. After testing whether the raw data fit the normal distribution or not by employing Shapiro-Wilks test, further statistical analyzes were performed with the most appropriate test parametric or non-parametric tests. Results: Statistically significant differences were found in terms of early apoptosis P0,01 and late apoptosis P0,001 cell population rates, while there was no significant difference of necrosis P0,064. Significant P0,001 increase in early apoptosis was evident in the majority of groups cultured differentiation medium for 48 or 72 hours, compared to the cells cultured with only HG-DMEM or differentiation media for 24 hours. While a significant increase in late apoptosis was observed in the majority of the groups cultured with the differentiation medium compared to the control group P0.001, no significant difference was observed between the groups differentiated for different durations P0,05. Conclusion: HT22 hippocampal cell line, which is frequently used in in-vitro modeling of important pathobiological features of neurodegenerative diseases, but poorly understood for the suitability in neuronal differentiation, has a markedly increased response to a widely used differentiation medium in terms of early apoptosis culturing duration dependent and late apoptosis regardless of duration. Our findings provide important clues at the level of apoptosis in terms of the usability of the HT22 cell line in differentiation studies.

 Amaç: Bu çalışmanın amacı nörobiyoloji çalışmalarında sıkça kullanılmakta olan HT22 fare hipokampal hücre hattının nöronal farklılaşma modeli olarak kullanılabilirliğinin anlaşılması için standart bir farklandırma besiyerine farklı sürelerde verdiği apoptotik cevabın sınanmasıdır. Gereç ve Yöntemler: HT22 hücrelerinin ekilmesi ve farklandırma dışında kültüre edilmesi için HG-DMEM farklandırma için ise B27 katkılı NB medyumu kullanılmıştır. Kontrol grubu da dâhil olmak üzere toplam 9 farklı grup standart olarak kültüre edilmiştir. Deney sonunda AnnexinVPI işaretlemesiyle erkengeç apoptozis ve nekrozis oranları akım sitometrik olarak belirlenmiştir. Verilerin normal dağılıma uyup uymadığı Shapiro-Wilks testi ile sınandıktan sonra en uygun testle istatistiksel analiz gerçekleştirilmiştir. Bulgular: Gruplar arasındaki karşılaştırmada erken apoptozis P0,01 ve geç apoptozis P0,001 hücre popülasyon oranları açısından istatistiksel olarak anlamlı bir fark tespit edilirken nekrozis açısından anlamlı bir fark olmadığı gözlenmiştir P0,064. 48 ve 72 saat farklandırma besiyerinde kültüre edilen grupların büyük çoğunluğunda, sadece HG-DMEM ile veya 24 saat farklandırılan hücrelere kıyasla istatistiksel olarak anlamlı düzeyde P0,001 erken apoptozis artışı olduğu görülmüştür. Farklandırma besiyeri ile kültüre edilen grupların çoğunluğunda kontrol grubuna kıyasla anlamlı düzeyde geç apoptozis artışı gözlenirken P0,001, farklı süreler farklandırılan gruplar arasında anlamlı bir fark gözlenmemiştir P0,05. Sonuç: Nörodejeneratif hastalıklara ait önemli patobiyolojik özelliklerin in-vitro düzeyde modellenmesinde sıkça kullanılan, fakat nöronal farklılaşma açısından uygunluğu yeterince bilinmeyen HT22 hippokampal hücre hattının, yaygın olarak kullanılan bir farklandırma besiyerine erken apoptozis kültürleme süresine bağlı ve geç apoptozis süreden bağımsız artışı şeklinde anlamlı tepkiler oluşturduğu anlaşılmıştır. Bulgularımız HT22 hücre hattının farklılaşma çalışmalarında kullanılabilirliği açısından apoptozis düzeyinde önemli ipuçları sunmaktadır.

Article Details

Measuring the Apoptotic Response of the HT22 Mouse Hippocampal Cell Line to Neuronal Differentiation Medium

Authors:

Ayla Batu Öztürk | Derya Yetkin | Nail Can Öztürk

Publisher:

Sakarya University

Main Topics:

Sağlık Kurumları Yönetimi

Journal:

Sakarya Tıp Dergisi Vol.13 Issue 3, pp.375-383

Document Type:

Article

Publication Date:

9/2023

Document Language:

Turkish

Document ID (Org No.):

2001298974

pISSN:

2146-409X

HT22

hippocampus

neuronal differentiation

apoptosis

flow cytometryHT22

hipokampus

nöronal farklanma

apoptozis

akım sitometri

Issues

Article Details

Authors: Ayla Batu Öztürk | Derya Yetkin | Nail Can Öztürk

Publisher: Sakarya University

Main Topics: Sağlık Kurumları Yönetimi

Journal: Sakarya Tıp Dergisi Vol.13 Issue 3, pp.375-383

Document Type: Article

Publication Date: 9/2023

Document Language: Turkish

Document ID (Org No.): 2001298974

pISSN: 2146-409X

Issues

Authors:

Ayla Batu Öztürk | Derya Yetkin | Nail Can Öztürk

Publisher:

Sakarya University

Main Topics:

Sağlık Kurumları Yönetimi

Journal:

Sakarya Tıp Dergisi Vol.13 Issue 3, pp.375-383

Document Type:

Article

Publication Date:

9/2023

Document Language:

Turkish

Document ID (Org No.):

2001298974

pISSN:

2146-409X