Tea Polyphenols as Natural Products for Potential Future Management of HIV Infection – An Overview

Belonging to the Lentivirus genus of animal retroviruses, human immunodeficiency virus (HIV) is the etiological agent of acquired immunodeficiency syndrome (AIDS) which attacks cells of the immune system including CD4+ T lymphocytes, monocytes, macrophages and dendritic cells. A rapid progression to immunodeficiency and the higher transmissibility of HIV-1 compared to HIV-2 are hallmarks of the worldwide spread of AIDS. Conventional HIV treatments are limited by drug toxicity and by multi-drug resistance, caused by the high genetic variability of HIV. This has led researchers into new areas of drug discovery in search of novel therapeutic molecules. Accumulating evidence indicates that tea polyphenols possess a range of beneficial properties including anti-cancer, anti-inflammatory, anti-oxidative, neuro-protective, antibacterial, anti-fungal and anti-viral effects. The anti-HIV infection potential of tea polyphenols has been confirmed by several preclinical studies. This suggests that polyphenol-rich extracts of tea could be used as dietary supplements as part of a combined therapeutic regimen with conventional anti-HIV drugs. Phenolic structures may also be considered as backbones for the discovery of a new generation of anti-HIV remedies. This review provides a perspective on the antiHIV activity of tea polyphenols and their development as a possible source of future drugs for the therapy of HIV/AIDS. Tea Polyphenols as Natural Products for Potential Future Management of HIV Infection – An Overview Hamid Reza Sodagari1, Roodabeh Bahramsoltani2, Mohammad Hosein Farzaei3, Amir Hossein Abdolghaffari4, Nima Rezaei5 and Andrew W. Taylor-Robinson6* 1Young Researchers & Elite Club, Karaj Branch, Islamic Azad University, Karaj, Iran 2 Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran 3Medical Biology Research Center, Kermanshah University of Medical Sciences, Kermanshah, Iran 4Pharmacology & Applied Medicine Department of Medicinal Plants Research Center, Institute of Medicinal Plants, ACECR, Karaj, Iran 5Research Center for Immunodeficiency, Children’s Medical Center, Tehran University of Medical Sciences, Tehran, Iran 6School of Medical & Applied Sciences, Central Queensland University, Rockhampton, Australia *Author for correspondence Email: a.taylor-robinson@cqu.edu.au JOURNAL OF NATURAL REMEDIES


Introduction
Ipomoea reniformis (IR) also called as merremia emarginata (Burm. f.) is a procumbent herb belonging to the family convolvulaceae. In India, it is commonly known as Undirkana and Mushakparni. The plant is widely distributed in India, Sri Lanka, Philippines, Malaysia, Tropical Africa and mainly grows in rainy and winter season. In India, it is found in Southern part mainly counting Chennai, and some places of Andhra Pradesh [1]. Traditionally, IR has been used to treat diverse clinical conditions ranging from pain; fever to neurological disorders [2]. IR has been claimed to be useful for inflammation, headache, fever, cough, neuralgia, rheumatism and also in liver and kidney diseases [3]. The powder of leaves is used as a snuff during epileptic seizures. Juice acts as purgative and the root is having diuretic, laxative actions and applied in the disease of the eyes and gums [4].
The plant contains various neuroprotective chemical constituents such as caffeic, p-coumaric, ferulic and sinapic acid esters. Petroleum ether extract contains fats and fixed oil while aqueous extract contains amino acids, tannins (condensed and pseudo tannins) and starch [5]. IR has been reported to possess various pharmacological actions, mainly antidiabetic [6], antiinflammatory [7], nephroprotective [8], antibacterial [9], antioxidant and antimicrobial activity [10]. Further, the principle constituents of IR such as sinapic and ferulic acids have exhibited behavioural and pharmacological

HIV Aetiology and Pathophysiology
Human Immunodeficiency Virus (HIV) is a member of the Lentivirus genus, part of the Retroviridae family of enveloped, single-stranded, positive-sense RNA viruses that are characterized by replication in a host cell through the process of reverse transcription. HIV is the etiological agent of Acquired Immuno Deficiency Syndrome (AIDS) which infects cells of the human immune system including CD4 + T lymphocytes, monocytes, macrophages and dendritic cells 1 . As a zoonotic disease HIV/AIDS is thought to be derived from simian immunodeficiency viruses that infect West African primates 2 . Common chimpanzees (Pan troglodytes) and western lowland gorillas (Gorilla gorilla gorilla), and sooty mangabey monkeys (Cercocebus atys), are recognized as the original source of HIV-1 and HIV-2, respectively. Of the two major types of HIV, such intrinsic features of HIV-1 as a greater rate of transmission than HIV-2 and driving a quicker loss of host immunocompetence have enabled it to become the principal cause of AIDS around the world 2 . M, N, O and P are the four identified groups of HIV-1 that are isolated from chimpanzees and gorillas. Of these, group M, which comprises nine subtypes: A-D, F-H, J and K, is known to be responsible for the global HIV pandemic. This group was transmitted to humans in West-Central Africa approximately 100 years ago 3,4 . The globalization of HIV-1 may be due to the high mutation and recombination rates of the nine subtypes of group M caused by the Reverse Transcriptase (RT) enzyme 5 . The number of infections caused by recombinants is reported to have increased from 17% over the period 2000-07 to at least 20% by 2011 6 . Drug injection with contaminated shared needles 7 , unprotected sexual contact, percutaneous blood exposure, mucous membrane exposure to contaminated blood or other body fluids, mother-to-child transmission and receipt of contaminated blood products are all identified as major routes of HIV transmission 8 . After gaining entry into the body via one of these routes, each virion attacks a CD4 + T lymphocyte, its main target, with the assistance of chemokine co-receptors such as CCR5 or CXCR4. The virus genome is released into the host cell and viral RT converts viral RNA into double stranded DNA. This new viral DNA is concealed within the DNA of the host cell by another HIV enzyme named integrase. In this way, transcription of the host cell's DNA during cell division causes spread of the virus within the host's body 9-11 .

HIV Epidemiology
It is estimated that worldwide over 35 million people live with HIV 12 , of whom 69% are resident in sub-Saharan Africa where 1 in every 20 adults is HIV-positive 13 . As far back as 2009 6% of Kenyan adults (15-49 years) were infected with HIV-1, with a difference between urban (10%) and rural areas (5.6%) 14

Current Treatment of AIDS Complications
There are more than 25 Anti-retroviral (ARV) drugs licensed for HIV therapy which are divided into several groups including nucleoside RT inhibitors (zidovudine, didanosine, zalcitabine, stavudine, lamivudine, abacavir, emtricitabine), nucleotide RT inhibitor (tenofovir), non-nucleoside RT inhibitors (nevirapine, delavirdine, efavirenz and etravirine), protease inhibitors (saquinavir, ritonavir, indinavir, nelfinavir, amprenavir, lopinavir, atazanavir, fosamprenavir, tipranavir and darunavir), fusion inhibitor (enfuvirtide), co-receptor inhibitor (maraviroc) and integrase inhibitor (raltegravir) 17 . Several side effects including constipation, fever, liver disorders, muscular dystrophy, lipoatrophy, metabolic disorders, blood disorders, hypersensitivity reactions, peripheral neuropathy, hyperlactaemia, pancreatitis and mitochondrial toxicity 1, 18,19 , as well as multi-drug resistance and toxicity caused by the high genetic variability of HIV have limited long term treatment with the aforementioned ARV drugs 20 . Thus, screening of alternative drugs such as plant metabolites used as natural products would appear necessary due to their therapeutic activities with lesser or no side effects. Against this background, the principal purpose of this review is to provide an overview of recent findings on anti-HIV activity of polyphenols extracted from different types of tea.

Health Benefits of Tea Polyphenols as Dietary Natural Products
Recently, polyphenols have been recognized as extremely important plant-derived dietary compounds due to producing a vast range of effects considered to be beneficial to humans. This comprises antioxidant, antibacterial, antiviral, anti-allergic, anti-colitis, anti-inflammatory, anti-arthritic and anti-cancer activities [21][22][23][24][25] . Among several polyphenol sources in the human diet, tea is one of the most consumed beverages around the world as a result of the pleasant taste, attractive aroma and health-promoting effects. Depending on the degree of processing and fermentation, tea is divided into green tea, which is consumed mostly in South East Asian countries, black tea, which is commonly consumed in the Indian subcontinent and Western societies, and Pu-erh, white, yellow and oolong teas that are produced and drunk mainly in China 26 . There are many reports about consumption of tea worldwide. The UK Tea Council claims that British people drink 60.2 billion cups of tea per year 27 . Worldwide, over 4.52 million tons of tea were produced in 2014 28 . In addition to the abundant distribution of polyphenols in this plant species, tea is also a leading natural source of caffeine, a psychoactive alkaloid that acts as a stimulant and, if taken at a bioactive dose, can show beneficial effects on wellbeing 29 . While used as a routine part of the human diet, tea has attracted scientific attention due mostly to its highly active antioxidant and anti-inflammatory polyphenolic compounds, especially catechins 29 . The European Food Safety Authority reported that each 100 ml of green tea contains about 126 mg of catechin derivatives 30 . Several beneficial properties have been attributed to tea polyphenols, including anti-cancer 31,32 , anti-diabetic 33 , anti-oxidative 34 , cardioprotective 35 and neuroprotective 36 activities, as well as antibacterial 37 , antifungal 38 and antiviral 39,40 functions. Considering the high content of antioxidants and anti-inflammatory components, including flavonoids, tea could be introduced as a suitable beverage for the prevention of chronic inflammatory and oxidative stress-related conditions 41 .
The immunoregulatory effects of green tea extracts have been reported [50][51][52][53] . A dose-dependent decrease in histamine production was noted for rodent peritoneal mast cells incubated with green tea polyphenol 50 . Further experiments by these authors showed that the altered histamine release was due to a green tea extract-mediated decrease in cAMP and calcium levels which leads to an NFκB and p38 MAPK-dependent inhibition of the proinflammatory cytokines TNF-α and IL-6 50 . Green tea components are reported to have immunoregulatory activity. EGCG was effective at suppressing peripheral blood mononuclear cell proliferation and IFN-γ production 51 . They can also change the profile of proinflammatory cytokines secreted by lymphocytes 52 . Neutrophils were modulated by green tea polyphenols through suppression of the TLR-4/NFκB p65 signal activation pathway 53 .
EGCG inhibits HIV-1 replication in several steps of the virus life cycle by interfering with RT and protease activity, blocking gp120-CD4 interaction via binding to CD4 and destroying virions 54- 58 59 . SEVI and also semen-mediated enhancement of HIV-1 infection may be inhibited by EGCG without cellular toxicity. In addition, the formation of fibril is abrogated completely by treatment with high concentrations of EGCG (10 and 20 mM EGCG) 60 . Furthermore, it has been reported that blockade of antibody and glycoprotein 120 through binding of EGCG to CD4 + T cell is another anti-HIV mechanism of this molecule 54 . Prevention of attachment of HIV-1glycoprotein 120 to CD4 has a key role in inhibition of HIV-1 infection by EGCG 61 . Also, inhibition of HIV-1 p24 antigen production across a broad spectrum of both HIV-1 clinical isolates and laboratory adapted subtypes (B, C, D and G) has been observed following application of EGCG at a concentration of 6 μmol/L. Inhibition of HIV-1 infection by EGCG is not a consequence of cytotoxicity, cell growth inhibition or apoptosis, indicating the specificity of the inhibitory potential of EGCG for HIV-1 infectivity 62 . Treatment of HIV-1 Tat transgenic mice with EGCG at a dose of 300mg/kg/day caused a reduction in Glial Fibrillary Acidic Protein (GFAP) associated neuronal loss 63 . EGCG both mildly reduced activated microgliosis and enhanced neuron survival. Antiviral activity of EGCG occurs by interacting with several steps of the HIV-1 life cycle. A destructive effect on virions, post-adsorption entry and RT in acutely infected monocytoid cells were also observed at concentrations of EGCG greater than 1 μM 56 . However, suppression of protease kinetics was identified at EGCG doses higher than 10 μM. Moreover, inhibition of virus production by THP-1 cells chronically infected with HIV-1 was promoted by liposome modification of EGCG in a dose-dependent manner. Following EGCG treatment of THP-1 cells inhibition of viral mRNA production was observed in lipopolysaccharide (LPS)activated chronically HIV-1-infected cells; however, inhibition of viral mRNA production in unstimulated or LPS-stimulated T-lymphoid cells (H9) was not observed 56 . Another in vitro study showed that EGCG strongly inhibited the replication of HIV as determined by RT and p24 assays performed on cell supernatants 55 . Among catechin derivatives including ECG, EGC, EGCG and Green Tea Extract (GTE) with HIV-1 RT inhibitory activity, EGCG and ECG were recognized as highly potent HIV-1 RT inhibitors 58 . In addition, reduction by EGCG of HIV neurotoxicity in the presence of IFN-γ via inhibition of Janus Activating Kinase/Signal Transducer and Activator of Transcription (JAK/STAT) pathway has been demonstrated in vivo 64 .

Pu-erh Tea Polyphenols
Pu-erh tea is a dark, aged tea that is produced in Yunnan province, China, from a large-leaved variety of tea plant, Camellia sinensis assamica, following a two-step process of microbial fermentation by organisms such as the mould Aspergillus sp. and oxidation 65,66 . Several beneficial activities of Pu-erh tea are documented, including prevention and treatment of cancer, heart diseases, rheumatoid arthritis and immune disease such as AIDS [67][68][69] . Due to its inhibitory effect on HIV replication Pu-erh tea extracts may be used as a dietary supplement or a natural product administered to HIV patients 65 . Water and ethanol extracts of Pu-erh tea (WEPT and EEPT), as well as fermented Pu-erh tea (FPT), possess anti-HIV activity with low cytotoxicity. The WEPT showed better anti-HIV activity than EEPT. Furthermore, this activity of the FPT's water extracts was better than that of the untreated Pu-erh tea. As shown in Table 1, it has been reported that HIV-1 RF and HIV-2 CBL-20 infections can be inhibited by Pu-erh tea extracts. Pu-erh tea reduced p24 antigen expression in HIV-1 IIIB acutely infected C8166 cells and HIV-1 KM018 infected peripheral blood mononuclear cells. The fusion between normal C8166 cells and HIV-1 chronically infected H9 cells was blocked by Pu-erh tea. The synergistic anti-HIV activity of Pu-erh tea in combination with the antiretroviral medication azidothymidine (AZT) was noted although no inhibitory effects on RT activity were observed 65 .

Black Tea Polyphenols
Chinese black tea is made from the small-leaved Camellia sinensis sinensis and is more oxidized and stronger in flavour compared to other varieties of tea. Theaflavins are polyphenols that are found naturally in black tea and which constitute approximately 2% of its dried water extract. Anti-bacterial, anti-viral, anti-inflammatory, anti-oxidative and anti-tumour activities have been reported as properties of major theaflavin derivatives in black tea including theaflavin (TF1), theaflavin-3-gallate (TF2A), theaflavin-3'-gallate (TF2B) and theaflavin-3,3'digallate (TF3) [70][71][72][73] . It is known that theaflavins in black tea possess anti-HIV-1 activity through inhibition of virus entry into target cells by interfering with glycoprotein 41 six-helix bundle formation 40 . Due to higher anti-HIV activity of black tea theaflavins in comparison to green tea catechins, extracts of the former are leading candidates for a new generation of anti-HIV-1 herbal therapy 74 . Theaflavins and catechins both inhibit HIV-1 entry by targeting glycoprotein 41 between the N-and C-peptides 40 . Amongst eight examined tea polyphenols, TF2B had the predominant anti-HIV activity followed in decreasing order by TF3, BEGCG, TH2A, EGCDG, GCG, TF1 and EGCG. Furthermore, the most inhibitory effect on HIV-1-mediated cell-cell fusion has been attributed to TF3 followed by GCG, TF2B, BEGCG, EGCDG, TH2A, TF1 and, poorly, EGCG. In contrast, neither inhibition of HIV-1-mediated cell-cell and virus-cell fusions, nor inhibitory activity against HIV-1 replication have been observed for some other tea polyphenols including (-)-catechin, CG, GC, EC, ECG and EGC 40 . It is thought that the mechanism of anti-HIV activity of tea polyphenols is not via specific interaction or interference with co-receptor binding. According to molecular docking analysis TF3 may bind to a highly conserved hydrophobic pocket that is located on the surface of the central trimeric coiled-coil of glycoprotein 41 40 . Another in vitro investigation using p24 production and luciferase assays demonstrated that a natural preparation containing 90% theaflavins (TFmix) had a strongly anti-HIV-1 effect on both laboratory-adapted and primary HIV-1 strains possessing low cytotoxicity 75 . Inhibition occurred at viral entry of the host cell by targeting glycoprotein 41 and blocking membrane fusion. While RT activity of the virus is also suppressed by TFmix, the IC 50 for this inhibition is approximately 8-fold higher than that needed for inhibition of glycoprotein 41 6-HB formation, thus indicating that RT is not a primary target for TFmix 74 . In an animal model of arthritis black tea also showed anti-inflammatory activity 76 , demonstrating the immunomodulatory properties of theaflavins, which may be considered as complementary to their apparent protective effect against HIV-1 discussed here.

Discussion
According to the World Health Organization, around 71 million persons have become infected with HIV since records began, of whom 34 million patients have died due to AIDS-related illnesses, with a further 36.9 million individuals identified as carrying HIV at the end of 2014 77 . Current medicaments for AIDS include nucleoside RT inhibitors, nucleotide RT inhibitor, protease inhibitors, fusion inhibitors, co-receptor inhibitors and integrase inhibitor 17 , but which in some patients may cause a range of adverse side effects like liver disorders, muscular dystrophy, metabolic disorders, blood disorders, hypersensitivity reactions, pancreatitis and mitochondrial toxicity 1, 18,19 . Multi-drug resistance and toxicity linked to the high genetic variability of the virus are additional limitations of conventional HIV treatments that collectively have prompted renewed efforts to discover novel therapeutic options.
Since ancient times humans have used plant-derived agents as treatments for various infective diseases and other ailments. The wealth of botanical-based therapies that are both efficacious and safe has prompted research to screen different known medicinal plants for anti-HIV activity, of which different types of tea -with their broad range of polyphenols as an active ingredientcould be considered as among the most important 41 . Green, Pu-erh and black teas contain such phenolic compounds as ECG, EGC, EGCG, GCG and TF which have demonstrated anti-HIV activity in several in vitro and in vivo studies (Table 1). While both are classified as flavonoids, green tea polyphenols consist mostly of EGCG and other catechin gallates, whereas theaflavins are the main constituents of black tea.
These two main categories of tea polyphenols are involved in cellular pathways such as JAK/STAT 64  were not assessed; however, these preparations were successful in demonstrating anti-HIV properties. Figure 1 illustrates the chemical structure of each of the tea polyphenols with anti-HIV activity most relevant to future drug preparation. As one of the most significant properties of polyphenols, anti-oxidant effects are likely to play a crucial role in their noted antiretroviral activities. In addition, an immunomodulatory effect could be one of the mechanisms involved in the anti-HIV function of these molecules.

Conclusion
Taking into account the collective body of research discussed herein, it is proposed that tea polyphenols and polyphenol-rich extracts could be used in future as dietary supplement in combination with conventional anti-HIV medicines. Phenolic structures may also be considered as scaffolds for the discovery of a new generation of anti-HIV drugs. Since investigations of the protective effects of specific tea polyphenols in humans are either limited or have not yet taken place, there is a pressing need to undertake detailed clinical trials in order to attain sufficient and robust data to either support or refute the predicted possible beneficial role of tea polyphenols in the therapy of HIV-infected patients.

Conflict of Interests
The authors do not have any conflict of interest to declare. This research received no specific grant from any funding agency in the public, commercial or notfor-profit sectors.