Nigella sativa amliorates inflammation and demyelination in the experimental autoimmune encephalomyelitis-induced Wistar rats

Abstract

Multiple sclerosis (MS) is the major, immune-mediated, demyelinating neurodegenerative disease of the central nervous system (CNS). Experimental autoimmune encephalomyelitis (EAE) is a well-established animal model of MS. The aim of the present study was to investigate the protective and ameliorative effects of N. sativa seeds (2.8 g/kg body weight) in EAE-induced Wistar rats. EAE-induced rats were divided into: 1- EAE-induced rats ("EAE" group). 2- "N. sativa + EAE" group received daily oral administration of N. sativa 2 weeks prior EAE induction until the end of the experiment. 3- "EAE + N. sativa" group received daily oral administration of N. sativa after the appearance of first clinical signs until the end of the experiment. All animals were decapitated at the 28th day post EAE-induction. EAE was investigated using histopathological, immunohistochemical and ultrastructural examinations in addition to determination of some oxidative stress parameters in the cerebellum and medulla. N. sativa suppressed inflammation observed in EAE-induced rats. In addition, N. sativa enhanced remyelination in the cerebellum. Moreover, N. sativa reduced the expression of transforming growth factor beta 1 (TGF β1). N. sativa seeds could provide a promising agent effective in both the protection and treatment of EAE.

Keywords: Experimental autoimmune encephalomyelitis (EAE); Nigella sativa (N. sativa); multiple sclerosis (MS); oxidative stress; transforming growth factor beta 1 (TGF-β1).

Figure 1

Effect of N. sativa administration on the cerebellar histopathological findings (H&E ×400). A. Control group showing the molecular layer containing a number of glia cells. B. Control group showing cerebellar medullary area containing glia cells. C. “EAE” group showing few lymphocytic infiltrations in the meningeal tissue associated with gliosis of the cerebellar molecular layer. D. “EAE” group showing reactive astrogliosis {note the astrocytes with eosinophilic increased cytoplasmic size (arrow head) and the appearance of axonal spheroid (arrow)}. E. “EAE + N. sativa” group showing mild gliosis of the cerebellar molecular layer with normal meningeal histology. F. “EAE + N. sativa” group showing mild reactive astrogliosis of the cerebellar medulla (arrow head). G. N. sativa + EAE” group showing mild gliosis of the cerebellar molecular layer with normal meningeal histology. H. “N. sativa + EAE” group showing mild astrogliosis in the cerebellar medulla that is only characterized by astrocytes hyperplasia not hypertrophy.

Figure 2

Effect of N. sativa administration on the brain stem histopathological findings (H&E ×400). A. Control group showing clusters of nerve nuclei surrounded by glia cells. B. Control group showing normal clear Virchow Robbin space. C. “EAE” group showing chromatolysis and necrosis of nerve nuclei with reactive astrogliosis (arrow). D. “EAE” group showing perivascular lymphocytic cuffing (arrow) with diffuse gliosis. E. “EAE + N. sativa” group showing necrosis of individual nerve nuclei (arrow). F. “EAE + N. sativa” group showing mild perivascualr mononuclear cell aggregation. G. “N. sativa + EAE” group showing gliosis with chromatolysis of individual nerve nuclei. H. “N. sativa + EAE” group showing focal gliosis of neuropill with scarce perivascular mononuclear cell aggregation.

Figure 3

Effect of N. sativa administration on the TGF-β1 expression in the rat brain. A. Control group showing negative immunolabeling astrocytes for TGF-β1 (×400). B. “EAE” group showing wide positive immunolabelling of astrocytes with strong positive brown color staining reaction (arrow) (×200), inserted box with higher magnification (×400). C. “EAE + N. sativa” group showing less and weak brown immunolabelling of astrocytes (×200). D. “N. sativa + EAE” group showing very few immunolabelling of astrocytes with very faint brown staining reaction (arrow) (×200), inserted box with higher magnification (×400).

Figure 4

Effect of N. sativa administration on the ultrastructure of myelin sheath in the rat medulla. (A) Control group, (B) “EAE” group, (C) “EAE + N. sativa” group and (D) “N. sativa + EAE” group. M: Myelinated fiber; T: Mitochondria; DF: Demyelinated fiber; PD: Partially demyelinated fiber; R: Remyelinated fiber; MO: Medium oligodendrocyte process; DO: Dark oligodendrocyte process; LA: Light astrocyte process and DA: Dark astrocyte process.

Figure 5

Effect of N. sativa administration on the ultrastructure of myelin sheath in the rat cerebellum. (A) Control group, (B) “EAE” group, (C) “EAE + N. sativa” group and (D) “N. sativa + EAE” group. M: Myelinated fiber; U: Unmyelinated fiber; T: Mitochondria; DF: Demyelinated fiber; R: Remyelinated fiber; MO: Medium oligodendrocyte process; DO: Dark oligodendrocyte process; LA: Light astrocyte process; DA: Dark astrocyte process and V: focal area of myelinolysis in the myelin sheath.