Pomegranate (Punica granatum L., Punicaceae) is used in the traditional medicine of different Asian cultures for the treatment of a variety of ailments. In Ayurvedic medicine, the plant, described under its Sanskrit name “dalima” (fruit), is considered as a “blood tonic” and used to cure parasitic infections . The decoction of the root was found beneficial in fevers and chronic debility due to malaria. Moreover, the fruit rind powder was found to possess immunomodulatory properties .
The eastern province of Orissa (India) is an area endemic for both Plasmodium falciparum and Plasmodium vivax; malaria constitutes a major health problem for the population, in particular for those living in rural areas. Since 1998, malaria patients referring to the Ayurveda dispensary receive a herbal preparation named OMARIA, made of sun-dried rind of the immature P. granatum fruits (Pg). OMARIA (the acronym for Orissa Malaria Research Indigenous Attempt) is distributed as a home based economic remedy for prophylaxis under the banner “Fight Malaria At Home/Ghare Maro Malaria. Clinical application started in 1998 by the Indian Red Cross Society Charitable Ayurveda dispensary (c/o District Magistrate Koraput) on behalf of D. Bhattacharya. Dispensary records indicate that OMARIA can successfully control P. falciparum and P. vivax infections in all patients including infants and pregnant women [3, 4]. OMARIA is administered as gelatine capsules (courtesy of m/s Sunil Health Care Ltd. New Delhi, India), containing each 825-850 mg of Pg. The therapeutic dose is one capsule every eight hours for three consecutive days. For prophylaxis, one capsule has to be taken in every day (children receive half the dosages) for a period ranging between two or four weeks/six months. Records from the Ayurveda-Indian Red Cross Society indicate a positive impact on the health status of the population under OMARIA coverage: the prophylactic intervention appears not only to reduce malaria episodes, but also the incidence of other infectious diseases, such as measles, chicken pox and conjunctivitis .
The reported anti-malarial effectiveness of the OMARIA was attributed to the anti-parasitic activity of a fraction enriched in tannins (Pg-FET) obtained from the Pg methanolic extract . The effect could be attributed to different constituents of Pg-FET, namely ellagic acid (EA), and punicalagin, which inhibited in vitro the growth of Pf asexual blood stages [5, 6]. Whether Pg preparations could help to control the malarial disease by adjuvant mechanisms, as well, remains unexplored. The present research was undertaken with the aim of testing the effects of Pg preparations on the pathways involved in the onset severe malaria, which may develops during Pf infection.
It is largely accepted that severe malaria is an inflammatory cytokine-driven disease. There is remarkable evidence that tumour necrosis factor (TNF) and interleukin-1 are important contributors to the systemic disease caused by the infectious agent Pf [7, 8].
The innate immune system seeks to destroy the protozoa and remove the remnants via phagocytosis by monocytes and neutrophils. Circulating levels of TNF, a mediator of the innate immune system, are increased after infection, as a consequence of stimulation of monocyte-macrophages by infected red blood cells (IRBC) or phagocytosis of haemozoin (Hz, the malarial pigment) phagocytosis by human monocytes [9–12]. TNF in turn enhances the synthesis of metalloproteinase-9 (MMP-9) in monocytes and macrophages [9, 10]. More recently, it has been shown that human monocytes fed with Hz or IRBC display increased MMP-9 activity and protein/mRNA expression and increased production of TNF and a role of MMP-9 and TNF in the onset of cerebral malaria has been postulated [11, 12].
The wide-range of therapeutic benefits of pomegranate have been attributed to its anti-oxidant and anti-inflammatory properties. It was shown that Pg fruit juice and its constituents ellagitannins, have a significant and broad inhibitory effect on MMPs, including MMP-9 [13, 14, 1]. In addition, Pg fruit juice and ellagitannins suppress inflammatory cell signalling induced by TNF in colon cancer cells .
However, it was not clear which of the compounds present in Pg-FET or ellagitannins could antagonize the host inflammatory response. In the present study, it has been investigated whether Pg-FET, EA, and punicalagin, the compounds previously identified in Pg-FET, inhibited MMP-9 upregulation and secretion in THP-1 cells induced by native haemozoin (Hz) and TNF. Since ellagitannins were shown to be converted by the human gut microflora into urolithin A, B and urolithin-8-methylether [16–20] these metabolites were also evaluated for their inhibitory effect.
Here, it is shown that Pg-FET and its constituents EA and punicalagin, all inhibited the MMP-9 secretion and expression in THP-1 cells, fed with Hz or TNF. Pg-FET and individual compounds were also able to inhibit MMP-9 promoter activity after stimulation with Hz. The inhibitory effect was partially due to the inhibition of NF-κB pathway. Urolithins were also active. It is then plausible that Pg, in addition to the direct effect on the parasite, modulates the malarial disease via the inhibition of the inflammatory response induced by haemozoin.