Objective: This study was aimed to evaluate the role of genistein or 4',5,7-trihydroxyisoflavone as a phytoestrogen in the treatment of estrogen deficiency-induced neuroinflammatory. The specific objectives of this study were to determine the anti-neuroinflammatory effect of genistein through measurement of MHC II and Arg1 expressions on microglia HMC3 cell line, as well as to prove that the effect occurs in an ER-dependent manner, through the measurement of free-ERβ expression. Methods: The cells were cultured in 24-well microplates, induced with 10 ng IFN-γ, and incubated for 24 h to activate the cell to M1 phenotype which has pro-inflammatory characteristics. Genistein with a concentration of 50 μM was added to the cells. The expression of MHC II, Arg1, and free-ERβ as markers was tested through an immunocytochemistry method and measured using the CLSM instrument. In silico approach was also conducted to determine the interaction between genistein and ERβ, compared to 17β-estradiol. Genistein structure was prepared with Avogadro 1.0.1, and molecular docking was done using PyRx 0.8 software. Biovia Discovery Studio Visualizer 2016 was used to visualize the structure of genistein against 3OLS protein. The physicochemical characteristics of genistein were analyzed using the SwissADME web tool. Results: Genistein can decrease MHC II expression and increase Arg1 expression in microglia HMC3 cells compared to negative controls (p<0.005), with expression value of 472.577±26.701 AU and 114.299±6.578 AU. But, genistein cannot decrease the free-ERβ expression in cells (p<0.005). The results of in silico analysis showed that genistein is an ERβ agonist. Conclusion: Genistein shows anti-neuroinflammatory effects by decreasing the MHC II expression and increasing Arg1 expression in the microglia HMC3 cell line. However, this effect does not occur through the binding of genistein to ERβ, but it is likely to occur through the binding of genistein with other types of ER.

Neurodegenerative disease is one of the problems faced by postmenopausal women due to estrogen deficiency. Phytoestrogen compounds can be used as an alternative treatment for diseases caused by estrogen deficiency by binding to their receptors through the estrogen receptor (ER) dependent pathway. With in silico studies, this study aims to predict how phytoestrogen compounds will stop neurons from dying by using the dependent ER pathway. Genistein, daidzein, glycitein, formononetin, biochanin A, equol, pinoresinol, 4-methoxypinoresinol, eudesmin, α-amyrin, and β-amyrin compounds were prepared with ChemDraw Ultra 12.0. Then their pharmacokinetic and pharmacodynamic properties were examined using SwissADME. Geometry optimization of the compound was performed using Avogadro 1.0.1, and molecular docking of the compound to the ERα (1A52) and ERβ (5TOA) receptors was performed using AutoDock vina (PyRx 0.8). The interaction visualization stage was carried out with Biovia Discover Studio 2021, while the toxicity values of the compounds were analyzed using pkCSM and ProTox II. The results showed that the equol compound met the pharmacokinetic, pharmacodynamic, toxicity criteria, and had similarities with the native ligand 17β-estradiol. Equol compound inhibits neurodegeneration via an ER-dependent pathway by binding to ERα (1A52) and ERβ (5TOA) receptors.

Neurodegenerative diseases are mostly experienced by postmenopausal women and are often caused by estrogen deficiency, so it is necessary to replace the hormone estrogen in the form of phytoestrogens. One of the plants that contain phytoestrogens is Marsilea crenata. The objective of this study is to see if giving rotenone-induced zebrafish a 96% ethanol extract of M. crenata leaves increases locomotor activity. This research was conducted by inducing 5 µg/L rotenone as a compound that will interfere with the locomotor activity of zebrafish. Next, treatment was given with 96% ethanol extract of clover in each group with a dose of 2.5; 5; 10; and 20 mg/mL to determine the effect of the extract on increasing locomotor activity of rotenone-induced zebrafish. Observations were made by looking at the quantity of zebrafish swimming every five minutes until day 28. The treatment of 96% ethanol extract of M. crenata leaves could significantly increase zebrafish motility at the optimum dose of 2.5 mg/mL, because every week the zebrafish locomotor activity increased compared to the negative control. M. crenata leaves extract is proven to prevent neurodegenerative diseases. However, further research needs to be done on the degenerative effects of rotenone every week.

Neuroinflammatory is a pathological condition in which neuronal cells experience progressive loss of function, one of which is due to estrogen deficiency. Phytoestrogens are compound that has structure and function similar to 17β-estradiol, the most potent natural form of estrogen. Semanggi (Marsilea crenata Presl.) is known to contain phytoestrogens which can replace the function of 17β-estradiol in maintaining organ homeostasis, so that is potential to be developed as an anti-neuroinflammatory drug. This study was aimed to prove the anti-neuroinflammatory effect of the n-hexane and ethyl acetate fraction of Semanggi leaves on HMC3 microglia cells, by measuring the expression of major histocompatibility complex II (MHC II). Both fractions with a concentration of 62.5; 125; and 250 μg/ml were administered to HMC3 microglia cells which had been induced by interferon gamma (IFN-γ) for 24 hours. Measurement of MHC II expression were using immunocytochemistry (ICC) methods and confocal laser scanning microscopy (CLSM) instruments. The results showed that n-hexane fraction could reduce MHC II expression at concentrations of 250 μg/ml with p<0.05, and showed a non-monotonic dose response (NMDR) pattern, while ethyl acetate fraction could reduce MHC II expression at all concentrations with p<0.05. Based on the results of the correlation test, the best anti-neuroinflammatory activity on HMC3 microglia cells is showed by the ethyl acetate fraction of Semanggi leaves.

Pendahuluan: Prevalensi penyakit neurodegeneratif meningkat tiap tahunnya dan mayoritas dialami perempuan yang mencapai fase menopause. Hal tersebut kemudian mengurangi kualitas hidup perempuan menopause. Penyakit neurodegeneratif pada wanita menopause sering diakibatkan oleh defisiensi estrogen, sehingga perlu adanya pengganti hormon estrogen dalam menjaga homeostasis sistem saraf pusat, salah satunya fitoestrogen. Salah satu tanaman yang mengandung fitoestrogen adalah semanggi (Marsilea crenata Presl.) Tujuan: Penelitian ini bertujuan mengetahui pengaruh pemberian ekstrak etanol daun M. crenata terhadap peningkatan aktivitas lokomotor ikan zebra. Metode: Penelitian ini dilakukan dengan induksi rotenon sebagai pengganggu aktivitas lokomotor ikan zebra, melalui pengukuran motilitas. Perlakuan dilakukan dengan dengan memberikan ekstrak etanol daun M. crenata pada dosis 2,5; 5; 10; dan 20 mg/mL untuk mengetahui pengaruh ekstrak terhadap peningkatan aktivitas lokomotor ikan zebra yang telah diinduksi rotenon. Pengamatan dilakukan dengan melihat kuantitas berenang ikan zebra setiap 5 menit pada hari ke 0; 7; 14; 21; dan 28. Kesimpulan: Pemberian ekstrak etanol 96% daun M. crenata dapat meningkatkan aktivitas lokomotor yang diketahui melalui peningkatan motilitas ikan zebra secara signifikan pada dosis optimum 2,5 mg/mL pada hampir tiap kelompok hari. Hal ini membuktikan ekstrak daun M. crenata berpotensi dalam mencegah progresifitas penyakit neurodegeneratif.