Network Analysis of Indigenous Indonesia Medical Plants for Treating Tuberculosis

Background: Indonesia is the biggest archipelago country with the second biggest biodiversity in the world. A lot of medical plants for treating various diseases can be found in Indonesia, including medical plants for tuberculosis, an infectious disease caused by Mycobacterium tuberculosis. Objective: The goal of this research is to document the information of Indonesia indigenous medical plants that used various local societies to treat tuberculosis and also analyze active compounds of medical plants with proteins that related to tuberculosis. Methods and Materials: The annotation of medical plants for treating tuberculosis was collected from a various source comprising local research papers, theses, and other resources. The information of active compound was taken from Dr. Duke’s Phytochemical and Ethnobotanical Databases. A network of active compounds-proteins was analyzed by using Cytoscape 3.6.0. Results: The result described that there were twenty-seven species from nineteen families of medical plants used by local societies of Indonesia for tuberculosis therapy, and there were sundry of active compounds from fourteen medical plants had direct interaction with proteins related tuberculosis. Conclusion: Most of the active compounds targeted proteins that had a prominent role in immune system. It indicated that medical plants treating tuberculosis through regulating immunity of human body.

7] Indonesia, a tropical archipelago country had vast biodiversity both natural and culture.A lot of indigenous medical plants grow in Indonesia, and local societies use it to treat a variety of diseases including tuberculosis.This study collected the information of medical plants used by local society of Indonesia to treat tuberculosis and analyze the involvement of active compounds with proteins related to tuberculosis by network analyzing.

Data Collection
In this study, various local resources like research papers, theses, and other resources were given ethnobotany information about the medicinal plants that used for treating tuberculosis in local society of Indonesia were collected.The data assembled were consisted of local name, the scientific name of the plants, location (Province), and part of the plants that used.The information of active compounds of the plants was obtained from Dr. Duke's Phytochemical and Ethnobotanical Databases (https:// phytochem.nal.usda.gov/phytochem/search).
This database provides not only about the active compound of the plants and the biological activity but also the information about the plant that commonly used for treating various diseases from around the world.Even there was a lot of information about ethnobotany in all of the countries, but unfortunately, this website gave limit information about the plant that used for tuberculosis in Indonesia local regions.

Network construction and analysis
Network analysis was used for understanding the effect of medical plants on tuberculosis.The network analyzing active compounds-proteins was constructed with string App of Cytoscape 3.6.0. 818 proteins related tuberculosis was obtained with STRING diseases feature and active compounds-proteins interaction was established with STITCH proteins/ compounds feature.4.0 cutoff score was used to take all of protein-protein and compounds-protein interaction.In the network graphic, proteins and active compounds were presented as nodes, while proteins-proteins and compounds-proteins interaction were presented as edges.

Plants used for treating tuberculosis in Indonesia Provinces
Through the literature retrieval, twenty-seven plants used local societies to treat tuberculosis from various provinces in Indonesia were obtained, as shown in Table 1.According to the Table 1, four species belong to Zingiberaceae, two species belong to Apiaceae, Malvaceae, Piperaceae, Euphorbitaceae and Rubiaceae, and one species respectively from Myrtaceae, Malvaceae, Fabaceae, Plantaginaceae, Piperaceae, Petiveriaceae, Lamiaceae, Rubiaceae, Verbenaceae, Euphorbiaceae, Apiaceae, Rutaceae, Moraceae, Acanthaceae, Bromeliaceae, Asphodelaceae, Asteraceae, and Araceae.According to Figure 1, Lantana camara L. and Curcuma domestica are precious tuberculosis medical plants for many local societies in Indonesia, followed by Centella asiatica, Hibiscus rosa sinensis, and Artocarpus elasticus.Lantana camara L is used extensively from west until east Indonesia provinces (map of Indonesia provinces is shown in Figure 2), 41 includes Lampung, Central Java West Sulawesi, and South Sulawesi, while   Curcuma domestica is most used only in East Indonesia Province such as East Java, Central Sulawesi, South Sulawesi, and East Kalimantan.Some of this medical plants not only in Indonesia but also in other countries also use it to treat tuberculosis.Leaves of Lantana camara L are used by local societies of Uganda to inhibit the activity of mycobacterial. 424] Mexican people use Citrus aurantifolia traditional medicine for tuberculosis, and moreover, it was already proved that Citrus aurantifolia peel could against multi-drug resistant Mycobacterium tuberculosis. 45Traditional China medicine plant, Zingiber officinallis Rosc.and Curcuma domestica are reported could medicate tuberculosis through isocitrate lyase and macrophage activity. 46  Tuberculosis therapies targeting immunity balancing can improve the treatment outcome and also well-regulated immune system may prevent reactivation of latent tuberculosis. 48The network describes some of the active compounds include ellagic acid, α-pinene, myristic acid, asiaticoside, aucubin, rutin, and esculin have direct interaction with protein related tuberculosis mechanism, while other compounds have indirect interaction.
Ellagic acid has direct interaction with IL-4, a cytokine produced by a variety of immune cells.In tuberculosis case, IL-4 has a role as an anti-inflammatory. 49However, The increasement of IL-4 was reported that could inhibit mycobacteria eradication through depletion of IFN-γ production. 50Ellagic acid, a phenolic compound found in a variety of plants including Euphorbia hirta.A previous study showed that ellagic acid could reduce the IL-4 level in eosinophilic inflammation case.Besides interacting with IL-4, ellagic acid also has interaction with Epigallocatechin gallate (EGCG) and NOS3 had a direct correlation with IL-4.In addition, Scoparone another active compound from Foeniculum vulgare is also targeting nitric oxide synthase 3 (NOS3), a macrophage enzyme produced nitric oxide that against microbial.NOS3 exhibit NO when Mycobacterium tuberculosis infects macrophage. 51sculin, one of an active compound found in Ocimum basillicum shows that interact directly with TNF, catalase (CAT) and Matrix metallopeptidase 9 (MMP9).It has been informed that TNF-α and MMP-9 had tuberculosis pathogenesis role.Mycobacterium tuberculosis through ERK pathway can elevate TNF-α and induce the production of MMP9. 526][57] In tuberculosis treatment, it may be suggested that esculin, gingerol, wogonin, and baicelin reduce the level of TNF-α and MMP9.Furthermore, zingerone found in Zingiber officinallis Rosc also have interaction with TNF-α through catalase.Catalase was stated that could induce apoptosis via TNF-α, which apoptosis for macrophage was an important mechanism to against mycobacterial infection. 58-59Prolyl 4-hydroxylase subunit beta (P4HB) is an enzyme catalyzing disulfide bonds that can increase Th-2 cells migration. 60P4HB is one of proteinrelated tuberculosis which targeted by rutin directly, whereas having indirect interaction with quercetin, luteolin, and curcumin through epidermal growth factor receptor (EGFR).2] In the network, the active compound of Plantago major and Centella asiatica, aucubin and asiaticoside, respectively can interact directly with IL-1B and CCL-2.Myristic acid and palmitate target TLR 4 which is related to tuberculosis pathogen.Toll-like receptor including TLR1, TKR2, TLR3, and TLR4, play a necessary part in the innate immune system.These receptors express in macrophage and dendritic cell to recognize mycobacterial.The recognition of TLR2 and TLR4 with Mycobacterium tuberculosis could induce macrophage apoptosis.In addition, palmitate can act as a TLR4 ligand on dendritic cells and induce IL-1B secretion. 63This may be specified that palmitate is a natural compound becoming a candidate for tuberculosis drug.CYP2B6 is one of cytochrome P450 enzyme involved in the transformation of drug and other xenobiotics, CYP2B6 polymorphism can be an indicator for tuberculosis treatment. 64α-pinene, a terpenoid compound, shows had direct interaction with CYP2B6.Even though other plants are not included in the network, but some previous studies reported the evidenced effect of tuberculosis.The ethyl-p-methoxycinnamate of Kaempferia galanga L can inhibit the activity of a variety of Mycobacterium tuberculosis strains including MDR strain. 65[68][69][70][71]

CONCLUSION
There are twenty-seven medical plants reported to treat tuberculosis disease in Indonesia local society.After being observed by network tuberculosis pathway analysis, there are some active compounds including ellagic acid, scoparone, esculin, zingerone, gingerol, baicalein, curcumin, rutin, quercetin, luteolin, asiaticoside, medacassoside, myristic acid, palmitate and α-pinene from fourteen plants such as Euphorbia hirta, Foeniculum vulgare, Ocimum basillicum, Zingiber officinallis Rosc, Curcuma domestica, Plantago major, Curcuma zedoaria, Centella asiatica, Coffea arabica, Ageratum conyzoides L, Tamarindus indica, Citrus aurantifolia, Petiveria alliacea and Lantana camara L that interact with protein related tuberculosis both directly and indirectly.Most of the active compounds target proteins involved in the immune system and it can be indicated that these compounds treat tuberculosis diseases through immune stability in the patient body.These plants may be a candidate to make a formulation for tuberculosis therapy and should be conducted in a real experiment.

Figure 1 :
Figure 1: Distribution of medical plants utilization used for tuberculosis.Figure 2: Map of Indonesia provinces.

Figure 2 :
Figure 1: Distribution of medical plants utilization used for tuberculosis.Figure 2: Map of Indonesia provinces.

Figure 3 .
Most of the active compounds targets are protein implicated in immune systems like IL-4, Tumor Necrosis Factor (TNF), IL-1B, CCL-2, and TLR4.It indicates that active compound treats tuberculosis through immunity balancing system.

Figure 3 :
Figure 3: Active compound-protein related tuberculosis pathway network.The red circle represents protein involved tuberculosis disease mechanism.The blue circle represents proteins which are not involved in tuberculosis mechanism.The yellow rectangular represent active compounds from medical plants.The green line represents active compound-protein interaction.The red line represents active compound-active compound interaction.The Grey line represent protein-protein interaction.
Bangladesh and Indonesia have similarity in medical plants for tuberculosis, it is reported that Andrographis paniculata, Centella asiatica, Aloe vera, and Hibiscus rosa sinensis are used to treat Mycobacterium tuberculosis infection.
47Analysis of active compounds target network Through Dr. Duke's Phytochemical and Ethnobotanical Databases active compounds of the medical plants were obtained from the database.In this study only selected active compounds were used, as shown in Table2.Based on the STITCH and STRING pathway analysis, it shows that several compounds from Euphorbia hirta, Foeniculum vulgare, Ocimum basillicum, Zingiber officinallis Rosc, Curcuma domestica, Plantago major, Curcuma zedoaria, Centella asiatica, Coffea arabica, Ageratum conyzoides L, Tamarindus indica, Citrus aurantifolia, Petiveria alliacea and Lantana camara L interact with protein related tuberculosis.The network constructed with Cytoscape is shown in