Comparison of Antioxidant mRNA expression levels of advanced Protruded and Extruded Nucleus Pulposus in Degenerative Lumbar Disc Herniations using the RT-PCR method

Year 2019, Volume: 5 Issue: 1, 1157 - 1167, 15.04.2019

Şeyho Cem Yücetaş , Necati Üçler

https://doi.org/10.30569/adiyamansaglik.486946

Abstract

Purpose: The aim of the present study is to compare the antioxidant levels of
nucleus pulposus in advanced protruded and extruded lumbar disc hernias (LDH)
with a steady lumbar posterior longitudinal ligament with the Real
Time-Polymerase Chain Reaction (RT-PCR) method.

Method: This
study was conducted with the comparison of the disc sections of a total of 40
patients who underwent an operation due to advanced protruded (n=20) and extruded
(n=20) LDH with the TRI-Reagent^® and Ribonucleic acid (RNA)
isolation and the RT-PCR method between January 2013 and May 2016. The study
included patients diagnosed with lumbar disc herniation who were applied with microdiscectomy
surgery. The antioxidant levels of the disc material, which caused compression
because it was highly protrudedorextruded, were detected using the RT-PCR
method,and the expression levels of the genes were determined.

Results: The antioxidant levels of the disc materials of a total of 40 patients, who
underwent an operation due to protruded and extruded disc hernia, were
determined with the RT-PCR method. The patients were found 25 females with a
mean age of 36.4 years and 15 males with a mean age of 39.26 years. Of the total
patients, 20 had MacNab grade 2 (protruded) and 20 had  MacNab grade 3 (extruded) disc hernias. The
expressions of protruded LDH materials were found to be lower than those of extruded
LDH materials.

Conclusion: Although the
expression levels of the significant antioxidant molecules of TGF-β, FGF-β,
IGF-1, NGF, MMP-3, and GAPDH mRNA, in patients with extruded LDH were
significantly higher compared to those with protruded LDH, no distinctive
features of these levels could be determined in terms of discogenic pain and
postoperative clinical improvement.

Keywords

Lumbar microdiscectomy, extruded disc, RT-PCR, Gene Expression

RT-PCR yöntemi kullanılarak Dejeneratif Lomber Disk Herniasyonunda İleri Düzey Protrüde ve Ekstrüde Nükleus Pulposusun Antioksidan mRNA ekspresyon düzeylerinin Karşılaştırılması

Abstract

Amaç. Bu
çalışmanın amacı, Real Time-Polimeraz Zincir Reaksiyonu (RT-PCR) yöntemi ile lumbar
posterior longitudinal ligamentte ileri düzey protrude ve ekstrüde lomber disk
hernilerinde (LDH) nükleus pulposusun antioksidan düzeylerini karşılaştırmaktır.

Metot: Bu
çalışma, Ocak 2013 ile Mayıs 2016 arasında izolasyon ve RT-PCR yöntemi ile ileri
düzey protrüde (n = 20) ve ekstrüde (n = 20) LDH ile TRI-Reagent® ve
Ribonükleik asit (RNA) ile ameliyat edilen toplam 40 hastanın disk bölümlerinin
karşılaştırılması ile yapıldı. Çalışmaya, mikrodisektomi cerrahisi uygulanan
lomber disk herniasyonu tanısı alan hastalar dahil edildi. Sıkışmaya neden olan
disk materyalinin antioksidan seviyeleri, RT-PCR yöntemi kullanılarak tespit
edildi ve genlerin ekspresyon seviyeleri belirlendi.

Bulgular: Protrude ve ekstrüde disk hernisi nedeniyle ameliyat edilen toplam 40
hastanın disk materyallerinin antioksidan düzeyleri RT-PCR yöntemi ile
belirlendi. Çalışmadaki 25 kadın hastanın yaş ortalaması 36.4 ve 15 erkek hastanın
yaş ortalaması 39.26 olduğu belirlendi. Toplamda hastaların 20'sinde MacNab
grade 2 (protruded) ve 20'sinde MacNab grade 3 (ekstrüde) disk fıtığı vardı. Ekspresyon
sonucunda  protrüde LDH materyallerinin
ekstrüde LDH materyallerinden daha düşük olduğu bulundu.

Sonuç: Ekstrüde
LDH'si olan hastalarda TGF-β, FGF-β, IGF-1, NGF, MMP-3 ve GAPDH mRNA'nın
anlamlı antioksidan moleküllerinin ekspresyon seviyeleri, protrüde LDH hastalara
göre anlamlı olarak daha yüksek bulunmasına rağmen, bu düzeylerin diskojenik
ağrı ve postoperative  klinik iyileşme
açısından ayırt edici özellik olarak belirlenemedi.

Keywords

Lomber mikrodiskektomi, ekstrüde disk, RT-PCR, Gen İfadesi

References

• 1. Jordan J, Konstantinou K, O’Dowd J. Herniated lumbar disc. BMJ Clin Evid. 2009 Mar;2009.
• 2. Sabnis AB, Diwan AD. The timing of surgery in lumbar disc prolapse: A systematic review. Indian J Orthop. 2014 Mar;48(2):127–35.
• 3. Schoenfeld AJ, Weiner BK. Treatment of lumbar disc herniation: Evidence-based practice. Int J Gen Med. 2010;3:209–14.
• 4. Soliman J, Harvey A, Howes G, Seibly J, Dossey J, Nardone E. Limited microdiscectomy for lumbar disk herniation: a retrospective long-term outcome analysis. J Spinal Disord Tech. 2014 Feb;27(1):E8–13.
• 5. Shamim MS, Parekh MA, Bari ME, Enam SA, Khursheed F. Microdiscectomy for lumbosacral disc herniation and frequency of failed disc surgery. World Neurosurg. 2010 Dec;74(6):611–6.
• 6. Wang D, Nasto LA, Roughley P, Leme AS, Houghton AM, Usas A, et al. Spine degeneration in a murine model of chronic human tobacco smokers. Osteoarthr Cartil. 2012;20(8):896–905.
• 7. Battie MC, Videman T, Kaprio J, Gibbons LE, Gill K, Manninen H, et al. The Twin Spine Study: contributions to a changing view of disc degeneration. Spine J. 2009;9(1):47–59.
• 8. Karabag H, Hatice S. The Relation of Lumbar Disc Herniation With Increased Lipid Hydroperoxide , Paraoxonase 1 and Total Oxidative Status. 2016;3(3):86–90.
• 9. Hou G, Lu H, Chen M, Yao H, Zhao H. Oxidative stress participates in age-related changes in rat lumbar intervertebral discs. Arch Gerontol Geriatr. 2014;59(3):665–9.
• 10. Chen JW, Ni B Bin, Li B, Yang YH, Jiang SD, Jiang LS. The responses of autophagy and apoptosis to oxidative stress in nucleus pulposus cells: Implications for disc degeneration. Cell Physiol Biochem. 2014;34(4):1175–89.
• 11. Wang H, Song Y, Cai L. Effect of percutaneous transforaminal lumbar spine endoscopic discectomy on lumbar disc herniation and its influence on indexes of oxidative stress. Biomed Res. 2017;28(21):9464–9.
• 12. Han Z, Wang J, Gao L, Wang Q, Wu J. Aberrantly expressed messenger RNAs and long noncoding RNAs in degenerative nucleus pulposus cells co-cultured with adipose-derived mesenchymal stem cells. Arthritis Res Ther [Internet]. 2018;20(1):182. Available from: https://arthritis-research.biomedcentral.com/articles/10.1186/s13075-018-1677-x
• 13. Silagi ES, Batista P, Shapiro IM, Risbud M V. Expression of Carbonic Anhydrase III, a Nucleus Pulposus Phenotypic Marker, is Hypoxia-responsive and Confers Protection from Oxidative Stress-induced Cell Death. Sci Rep [Internet]. 2018;8(1):1–13. Available from: http://dx.doi.org/10.1038/s41598-018-23196-7
• 14. Thermo Scientific molecular biology workflow solutions Supporting great science through innovation in molecular biology.
• 15. Tawa N, Rhoda A, Diener I. Accuracy of magnetic resonance imaging in detecting lumbo-sacral nerve root compromise: a systematic literature review. BMC Musculoskelet Disord. 2016;17(1):386.
• 16. Wassenaar M, van Rijn RM, van Tulder MW, Verhagen AP, van der Windt DAWM, Koes BW, et al. Magnetic resonance imaging for diagnosing lumbar spinal pathology in adult patients with low back pain or sciatica: a diagnostic systematic review. Eur spine J Off Publ Eur Spine Soc Eur Spinal Deform Soc Eur Sect Cerv Spine Res Soc. 2012 Feb;21(2):220–7.
• 17. Wiesel SW, Weinstein JN HH et al. The Lumbar Spine. 2nd ed. Philadelphia: WB Saunders; 1996.
• 18. Haro H, Crawford HC, Fingleton B, MacDougall JR, Shinomiya K, Spengler DM, et al. Matrix metalloproteinase-3-dependent generation of a macrophage chemoattractant in a model of herniated disc resorption. J Clin Invest. 2000 Jan;105(2):133–41.
• 19. Haro H, Komori H, Kato T, Hara Y, Tagawa M, Shinomiya K, et al. Experimental studies on the effects of recombinant human matrix metalloproteinases on herniated disc tissues--how to facilitate the natural resorption process of herniated discs. J Orthop Res. 2005 Mar;23(2):412–9.
• 20. Kim H-J, Park J-B, Won H-Y, Chang H. Serum Levels of TGF-β1, TIMP-1 and TIMP-2 in Patients with Lumbar Spinal Stenosis and Disc Herniation. Asian Spine J [Internet]. 2007;1(1):8. Available from: http://asianspinejournal.org/journal/view.php?doi=10.4184/asj.2007.1.1.8
• 21. Tolonen J, Grönblad M, Vanharanta H, Virri J, Guyer RD, Rytömaa T, et al. Growth factor expression in degenerated intervertebral disc tissue. An immunohistochemical analysis of transforming growth factor beta, fibroblast growth factor and platelet-derived growth factor. Eur spine J Off Publ Eur Spine Soc Eur Spinal Deform Soc Eur Sect Cerv Spine Res Soc. 2006 May;15(5):588–96.
• 22. Aoki Y, Nakajima A, Ohtori S, Takahashi H, Watanabe F, Sonobe M, et al. Increase of nerve growth factor levels in the human herniated intervertebral disc: Can annular rupture trigger discogenic back pain? Arthritis Res Ther. 2014;16(4):4–5.