title: In vitro and In vivo antimycobacterial, hepatoprotective and immunomodulatory activity of Euclea natalensis and its mode of action.
abstract: BACKGROUND
  The Natal gwarri or Natal ebony (Euclea natalensis A.DC.) is a deciduous tree found widespread throughout southern Africa, especially in Kwazulu-Natal and the southern cost. It has been widely used by indigenous communities such as the Zulus, Tsongas and Vendas for symptoms related to tuberculosis (TB). The decoctions made from the plant parts are administered for chest diseases to treat complications such as chest pains, bronchitis, pleurisy and asthma. TB is prevalent in immune-compromised patients and it is evident that TB-drugs cause hepatotoxicity. The objective of the present study was therefore to evaluate the antimycobacterial activity of the ethanolic extract of E. natalensis against TB and its hepatoprotective and immunomodulatory activities.
  
  
  METHODS
  The antimycobacterial, antioxidant, hepatoprotective, immunomodulatory activity and cytotoxicity of the ethanolic extract of the shoots of E. natalensis were determined in vitro. The mechanism of action of the antituberculosis activity was determined by investigating the inhibitory effect on mycothiol disulfide reductase enzyme. Furthermore, the acute, sub-acute toxicity (50-2000mg/kg) and antimycobacterial effect (300mg/kg) of E. natalensis shoot extract were investigated in Balb/c mice. Hepatoprotective activity of the extract (50-150mg/kg) was evaluated on isoniazid and rifampicin (50mg/kg; i.p.) induced hepatic damage in a rat model.
  
  
  RESULTS
  The minimum inhibitory concentration of the extract was found to be 125µg/ml against Mycobacterium tuberculosis. The extracts 50% inhibitory concentration (IC50) against 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical was found to be 22.55µg/ml. The plant showed a hepatoprotective effect (50% at 12.5µg/ml) and the ability to increase T-helper 1 cell cytokines; Interleukin 12, Interleukin 2 and Interferon α by up to 12 fold and the ability to decrease the T-helper 2 cell cytokine Interleukin 10 4 fold when compared to baseline cytokine production. No cellular toxicity was observed in primary peripheral blood mononuclear cells (PBMC's) and two secondary cell lines; U937 monocytes and Chang liver cells (a derivative of the HepG2 cell line). During mechanistic studies, the extract showed a 50% inhibition of mycothiol reductase activity at 38.62µg/ml. During the acute and sub-acute studies, E. natalensis exhibited no toxic effect and the 50% lethal dose (LD50) was established to be above 2000mg/kg. The extract was able to reduce the mycobacterial load (1.5-fold reduction) in infected mice. Isoniazid and rifampicin caused significant hepatic damage in rats, and the extract was able to reduce the toxicity by 15% and 40% at 50 and 150mg/kg respectively.
  
  
  CONCLUSIONS
  The present study supports the traditional usage of the plant against tuberculosis symptoms. The study showed the ability of E. natalensis shoot extract to inhibit mycobacterial growth, stimulate an appropriate immune response and have a hepatic protective effect. Due to the extract's significant results for hepatoprotective, immunomodulatory effects and antimycobacterial activity, it may prove to be effective to serve as an adjuvant for TB-patients.
authors: Department of Plant and Soil Sciences, University of Pretoria, South Africa. Electronic address: namrita.lall@up.ac.za.
authors: Namrita
authors: N
authors: Lall
authors: Department of Plant and Soil Sciences, University of Pretoria, South Africa. Electronic address: vivek494@gmail.com.
authors: Vivek
authors: V
authors: Kumar
authors: Department of Plant and Soil Sciences, University of Pretoria, South Africa; Faculty of Science, University of Johannesburg, South Africa. Electronic address: dmeyer@uj.ac.za.
authors: Debra
authors: D
authors: Meyer
authors: Department of Plant and Soil Sciences, University of Pretoria, South Africa. Electronic address: nomasomi.gasa@gmail.com.
authors: Nomasomi
authors: N
authors: Gasa
authors: Department of Plant and Soil Sciences, University of Pretoria, South Africa; Department of Pharmacy, University of East Anglia, Norwich, United kingdom. Electronic address: C.Hamilton@uea.ac.uk.
authors: Chris
authors: C
authors: Hamilton
authors: Department of Pharmacology, University of the Free State, South Africa. Electronic address: MatsabisaMG@ufs.ac.za.
authors: Motlalepula
authors: M
authors: Matsabisa
authors: Department of Plant and Soil Sciences, University of Pretoria, South Africa. Electronic address: u04405765@tuks.co.za.
authors: Carel
authors: C
authors: Oosthuizen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D000995:Antitubercular Agents / Q000494:pharmacology*; D002460:Cell Line; D031262:Ebenaceae / Q000737:chemistry*; D005260:Female; D006801:Humans; D007155:Immunologic Factors / Q000494:pharmacology*; D066298:In Vitro Techniques; D008099:Liver / Q000187:drug effects*; D042965:Methylenetetrahydrofolate Reductase (NADPH2); D051379:Mice; D008807:Mice, Inbred BALB C; D062105:Molecular Docking Simulation; D010936:Plant Extracts / Q000494:pharmacology*; D017386:Sequence Homology, Amino Acid
chemical_list: D000975:Antioxidants; D000995:Antitubercular Agents; D007155:Immunologic Factors; D010936:Plant Extracts; D042965:Methylenetetrahydrofolate Reductase (NADPH2)
keywords: Antimycobacterial; Euclea natalensis; Hepatoprotection; Immunomodulation; Toxicity