Sains Malaysiana 48(9)(2019): 1937–1946

http://dx.doi.org/10.17576/jsm-2019-4809-15

Effect of N-Acetylcysteine Supplementation on Oxidative Stress-Mediated Cryoinjury of Bone Marrow Derived-Hematopoietic Stem Cells

(Kesan Suplementasi N-Asetilsistein ke atas Kecederaan Krio Aruhan Tekanan Oksidatif pada Sel Tunjang Hematopoetik Pencilan Sumsum Tulang)

SHAWAL MARADONA ABDUL WAHAB, ZARIYANTEY ABD HAMID*, RAMYA DEWI MATHIALAGAN & IZATUS SHIMA TAIB

Biomedical Science Programme, Centre for Health & Applied Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Federal Territory, Malaysia

Received: 18 February 2019/Accepted: 19 June 2019

ABSTRACT

Hematopoietic stem cells (HSCs) transplantation was introduced as curative treatment for various diseases. Cryopreservation of HSCs is crucial for long term storage and maintenance of cellular quality; however, it has been reported that cryopreservation itself causes oxidative stress-driven apoptosis and cell loss. This study investigated impact of supplementing N-acetylcysteine (NAC) as antioxidant during cryopreservation on viability and oxidative stress in HSCs. HSCs were isolated from murine bone marrow, cultured in HSCs growth media and cryopreserved (1×106 cells per vial) together with 10% DMSO and NAC (0, 0.25, 0.5 or 2.0 μM) for 48 h, 2 weeks or 8 weeks at -196°C using controlled-rate-freezing technique. Cell viability and oxidative stress in cryopreserved cells were analysed at each time-point. Cell viability was significantly reduced (p<0.05) following cryopreservation as compared to pre-cryopreservation. NAC supplementation significantly increased cell viability (p<0.05) after 48 h cryopreservation at 0.5 μM and 2.0 μM and after 2 weeks cryopreservation at 0.25 μM compared to the controls. Cryopreservation significantly enhanced GSH level (p<0.05) and reduced MDA level (p<0.05) without affecting SOD activity and PC level in HSCs compared to pre-cryopreservation. NAC supplementation significantly increased GSH level at 0.25 μM in cryopreserved HSCs compared to control. In conclusion, NAC supplementation during cryopreservation showed potential in minimizing cryoinjury by promoting cell viability, increasing antioxidant capacity and reducing oxidative damage in HSCs, however these effects are influenced by both durations of cryopreservation and NAC concentration.

Keywords: Cryopreservation; hematopoietic stem cells; N-acetylcysteine; oxidative stress

ABSTRAK

Pemindahan sel stem hematopoietik (SSH) telah diperkenalkan sebagai rawatan pelbagai penyakit. Pengawetan krio SSH adalah penting untuk penyimpanan dan pemeliharaan kualiti SSH, namun pengawetan krio dilaporkan berupaya mengaruh apoptosis dan penurunan keviabelan sel akibat tekanan oksidatif. Kajian ini telah mengkaji kesan suplementasi N-asetilsistien (NAC) sebagai antioksidan semasa pengawetan krio terhadap keviabelan dan tekanan oksidatif SSH. SSH diisolasi daripada sum-sum tulang mencit, dikulturkan di dalam media pengkulturan SSH dan seterusnya diawet krio (1×106 sel per vial) dengan penambahan 10% DMSO dan NAC (0 μM, 0.25 μM, 0.5 μM atau 2.0 μM) selama 48 jam, 2 minggu atau 8 minggu pada suhu -196°C. Keviabelan sel dan tekanan oksidatif ditentukan ke atas SSH pada setiap tempoh pengawetan krio. Keviabelan sel menurun secara signifikan (p<0.05) selepas pengawetan krio berbanding dengan pra-pengawetan krio. Suplementasi NAC telah meningkatkan keviabelan sel secara ketara (p<0.05) selepas pengawetan krio selama 48 jam pada kepekatan 0.5 μM dan 2.0 μM dan juga selepas 2 minggu pengawetan krio pada kepekatan 0.25 μM berbanding kawalan. Pengawetan krio memberi kesan peningkatan aras GSH (p<0.05) dan penurunan aras MDA (p<0.05) tanpa memberi kesan yang signifikan terhadap aras SOD dan PC berbanding pra-pengawetan krio. Suplementasi NAC meningkatkan aras GSH secara signifikan (p<0.05) pada kepekatan 0.25 μM dalam SSH yang dikrio awet berbanding kawalan. Secara kesimpulannya, suplementasi NAC semasa pengawetan krio berpotensi meningkatkan keviabelan sel dan kapasiti antioksidan serta mengurangkan kesan kerosakan oksidatif, tetapi kesan ini adalah dipengaruhi oleh jangka masa pengawetan krio dan kepekatan NAC yang digunakan.

Kata kunci: N-asetilsistien; pengawetan krio; sel stem hematopoietik; tekanan oksidatif

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*Corresponding author; email: zyantey@ukm.edu.my