The effect of Ethyl Acetate and N-butanol Fractions of Extract of Arimonia eupatoria on the Repair of Spinal Cord and the Expression of Caspase 3 and 9 genes after Sciatic Nerve Compression in Rats
Subject Areas : Journal of Animal BiologyGhazale Nabavinia 1 , Maryam Tehranipour 2 , javad Baharara 3
1 - Department of Biology, Islamic Azad University, Mashhad Branch, Mashhad, Iran
2 - Department of Biology, Islamic Azad University, Mashhad Branch, Mashhad, Iran
3 - Department of Biology, Islamic Azad University, Mashhad Branch, Mashhad, Iran
Keywords: Agrimonia eupatoria, Sciatic nerve repair, Caspase 3 and 9,
Abstract :
Caspase 3 gene plays a role in the progression of degeneration and caspase 9 in apoptotic, necrotic, and lysosomal deaths, and after the nerve damage, This study aimed to determine the neuroprotective effects of hydro alcoholic extract and fractions of Agrimonia eupatoria plant on neuron density and the change in the expression of caspase 3 and 9 genes 36 male Wister rats weighing were divided to 6 groups including control, compression, treatment A (compression + hydro alcoholic extract in dose of 75 mg/kg), treatment B (compression + ethyl acetate fraction in dose of 75 mg/kg dose), Treatment C (compression +N butanol fraction in dose of 75 mg/kg) and treatment D (compression + aqueous phase with dose of 75 mg/kg). In the control group, after anesthetizing of the rats, the muscle at the sciatic nerve site was split without nerve damage and in compression group and treatment groups; sciatic nerve of the right leg was subjected to compression for thirty seconds. The first intraperitoneal injection of the extract was performed immediately after compression of the nerve and the second injection was performed 7 days later. After twenty-eight days, the rats were subjected to the perfusion method and samples were taken from the lumbar spinal cord. Slides were stained with toluidine blue, the neuron density was measured using a dissector method and the stereology method and the results were compared by t-test and Anova analysis. Spinal cord samples were taken, Total RNA extracted, and cDNA was synthesized, and then, and the expression changes of caspase 3 and 9 genes in the samples were investigated. The results showed that neuronal density increased significantly in all treatment groups compared to the compression group (p < 0.001) and it is more in the n-butanol group than the other groups. The gene expression of Caspase 3 and 9 decreased in all treatment groups compared with compression group (p < 0.01). The extract of this plant can be effective in repairing the nervous system due to the presence of compounds such as tocopherol, which has anti-oxidant and anti-apoptotic effects.
1. Adhiah A.H., Al-bederi O., Al-sammarrae K.W. 2013. Cytotoxic effects of Agrimonia eupatoria L. against.cancer cell lines in vitro. J Assoc Arab Univ Basic Appl Sci, 14: 87-92, 2013.
2.Al-snafi A. 2015. AE: The pharmacological and therapeutic importance of Agrimonia eupatoria - a review. Asian J Pharm Sci. Technol, 5: 112–117.
3. Bae H., Kim H-J., Shin M., Lee H., Yin C.S., Ra J., Kim J. 2010. Inhibitory effect of Agrimoniae Herba on lipopolysaccharide-induced nitric oxide and proinflammatory cytokine production in BV2 microglial cells. Neurol Res, 32: 53–57.
4. Behnam-Rasouli M., Nikravesh MR., Mahdavi-Shahri N. and Tehranipour M. 2000. Post-Operative time effects after sciatic nerve crush on the number of alpha motoneurons ,using a stereological (Disector). Iran. Biomed. J, 4(1): 45-9.
5. Benchoua A., Couriaud C., Guegan C. 2002.Active caspase-8 trans-locates to the nucleus of apoptotic cells to inactivate poly(ADP-ribose) polymerase-2. J Biol Chem, 277:34217–22.
6. Cain K., Bratton SB., Cohen GM. 2002. The Apaf-1 apoptosome: A large caspase-activating complex. Biochemie, 84:203–14.
7. Cande C., Cohen I., Daugas E. 2002. Apoptosis-inducing factor (AIF): A novel caspase-independent death effector released from mitochondria. Biochimie, 84:215–22.
8. Correia H.S., Batista M.T., Dinis T.C.P. 2007. The activity of an extract and fraction of Agrimonia eupatoria L. against reactive species. BioFactors, 29: 91–104
9. Ferrer I., Planas A.M. 2003. Signaling of cell death and cell survival following focal cerebral ischemia: life and death struggle in the penumbra. J Neuropathol Exp Neurol, 62(4): 329-39.
10.Gyrd-Hansen M., Farkas T., Fehrenbacher N., Bastholm L., Høyer-Hansen M., Elling F., Wallach D., Flavell R., Kroemer G., Nylandsted J., Jäättelä M. 2006. Apoptosome-independent activation of the lysosomal cell death pathway by caspase-9. Mol Cell Biol, 26(21):7880-91.
11. Hernando R. 2010. Neurological improvement of injured sciatic nerve following omental transplantation. J Neurol Sci , 27(2): 254-6.
12. Leker R.R., Shohami E. 2002.Cerebral ischemia and trauma—Different etiologies yet similar mechanisms: Neuroprotective opportunities. Brain Res Rev, 39:55–73.
13. Nesic O., Xu GY., McAdoo D., High K.W., Hulsebosch C. and Perez-Pol R. 2001. IL-1 receptor antagonist prevents apoptosis and caspase-3 activation after spinal cord injury. J Neurotrauma, 18(9): 947-56.
14.Rakhshandah H., Shakeri MT. and Ghasemzadeh MR. 2007. Comparative Hypnotic Effect of Rosa damascena Fractions and Diazepam in Mice. Iranian J Pharmaceuti Res, 6(3): 193-197.
15.Sommer C., Schomacher M., Berger C., Kuhnert K., Muller H.D., Schwab S., Schabitz W.R. 2006. Neuroprotective cannabinoid receptor antagonist SR141716A prevents downregulation of excitotoxic NMDA receptors in the ischemic penumbra. Acta Neuropathol, 112(3):277-86.
16. Strasser A., O’Connor L., Dixit V.M. Apopotosis signaling. 2000. 69:217–45.
17. Sun X.M., Bratton S.B., Butterworth M., MacFarlane M., Cohen G.M. 2002. Bcl-2 and Bcl-xL inhibit CD95-mediated apoptosis by preventing mitochondrial release of Smac/DIABLO and subsequent inactiva-tion of X-linked inhibitor-of-apoptosis protein. J Biol Chem, 277:1345–51.
18. Tehranipour M. Javadmoosavi Z. 2011. The neuroprotective effect of alcoholic extract of cannabis sativa on neuronal density of spinal cord alpha motoneurons after sciatic nerve injury in rat .j shaheed sadoughi uni med sci, 19(3). 49-339.
19. Tehranipour M. and Ghadamyari T. 2010. The effects of root aquatic extract of Salvia staminea on neuronal density of alpha motoneurons in spinal cord anterior horn after sciatic nerve compression in rat. J Biol Sci, 10(1): 48-52.
20. Zoltan P. , Biriczova L. , Pallag G., Carvalheiro Marqes E., Vargova N., Kmonickova E.2020. The Therapeutic Effects of Agrimonia eupatoria L. Physiol. Res, 69(4):S555-S571