Multi-omic analysis revealed the therapeutic mechanisms of Alpinia oxyphylla fructus water extract against bladder overactivity in spontaneously hypertensive rats.

OBJECTIVE: Alpinia oxyphylla fructus without impurities and shells is called "Yi-Zhi-Ren" (YZR) in Chinese, and traditionally used to alleviate enuresis. The aim of this study was to investigate the effects and underlying mechanisms of YZR in the treatment of overactive bladder (OAB) in spontaneously hypertensive rats (SHR), a vascular disorder-related OAB model. METHODS: A 3-week administration of YZR water extract (p.o.) was done, followed by urodynamics to measure bladder parameters. Changes in bladder structure were observed through H&E staining and Masson's staining. An integrated approach involving network pharmacology, transcriptomics and metabolomics was employed to elucidate the potential mechanisms of YZR, and the key proteins involved in the mechanisms were validated by Western blotting. Additionally, network pharmacology was used to predict the relationship between YZR's active components and validated proteins. RESULTS: YZR treatment significantly improved the bladder storage parameters, tightened the detrusor layer, reduced inflammatory infiltration, and decreased collagen proportion in the SHR bladder. These results indicated that YZR water extract can alleviate OAB symptoms and improve bladder structure. Integrated analysis suggested that YZR may affect extracellular matrix-receptor interaction and calcium signaling pathway. Western blotting results further confirmed that the reduction in key proteins, such as TGFß1, p-SMAD3, collagen III, Gq and PLCß1, involved in collagen synthesis and calcium signaling pathways after YZR treatment. Network pharmacology predicted that sitosterol, chrysin, and nootkatone were potential components responsible for YZR's therapeutic effect on OAB. CONCLUSION: YZR's mechanisms of action in treating OAB involved the TGFß1-SMAD3 signaling pathway-related collagen synthesis and Gq-PLCß1 calcium signaling pathway, which are associated with detrusor contraction frequency and strength, respectively.

Multi-omic approaches provide insights into the molecular mechanisms of Sojae semen germinatum water extract against overactive bladder.

Sojae semen germinatum (SSG) is derived from mature soybean seeds that have been germinated and dried, typically with sprouts measuring approximately 0.5 cm in length. SSG is traditionally known for its properties in clearing heat and moisture. Nevertheless, limited information was reported on the effects and mechanisms of SSG in alleviating urinary symptoms. This study employed urodynamic parameters to investigate the therapeutic effect of SSG water extract on overactive bladder (OAB) in the rat model with benign prostatic hyperplasia. Through a combination of transcriptomic and metabolomic analyses, the pathways and key proteins of the SSG treatment for OAB were identified and validated by ELISA and Western blotting. Furthermore, network pharmacology elucidated the roles of SSG's isoflavones acting on the target which was identified by above-mentioned multi-omics analysis. Our results indicate that SSG water extract significantly mitigated OAB by down-regulating the PGE2/EP1/PLCß2/p-MLC signaling pathway. It was speculated that the active ingredient in the SSG on EP1 was genistein. This study provided valuable insights into the molecular mechanisms of SSG water extract, emphasizing the multi-target characteristics and critical pathways in improving OAB. Furthermore, this study contributes to the potential utilization of SSG as a functional food.

Daidzein is the in vivo active compound of Puerariae Lobatae Radix water extract for muscarinic receptor-3 inhibition against overactive bladder.

Background: In the previous study, Puerariae Lobatae Radix (named Gegen in Chinese) water extract attenuated M3 receptor agonist carbachol-induced detrusor contraction after 3-week oral administration in a hypertension-associated OAB (overactive bladder) model. This research aimed to investigate the active ingredients from Gegen water extract against OAB. Methods: Bioassay-guided fractionation was performed by using preparative HPLC for fast isolation of fractions followed by screening their ex vivo activity through carbachol-induced bladder strip contraction assay. Chemicals in each active fraction were analyzed by HPLC-UV. Urine metabolites were quantified by LC-MS/MS after sub-acute administration. Thermal shift assay with the recombinant human M3 receptor protein was performed, and molecular docking analysis was used for molecular modelling of M3 receptor inhibition. Results: Bioassay-guided fractionation results for isolating M3 receptor inhibitors indicated that four compounds were identified as active ingredients of Gegen water extract, and their inhibition potency on carbachol-induced detrusor contraction was ranked in descending order according to their inhibition concentrations as follows: genistein > daidzein > biochanin A >> puerarin. Daidzein in urine reached an ex vivo effective concentration to inhibit detrusor contraction, but others did not. Daidzein concentration-dependently increased the melt temperature (Tm) of recombinant human M3 receptor protein with a positive binding (ΔTm = 2.12 °C at 100 µg/ml). Molecular docking analysis showed that daidzein can potently bind to the ligand binding pocket of the M3 receptor via hydrogen bonding. Conclusion: Puerarin and its derivatives were pro-drugs, and daidzein was their in vivo active form via M3 receptor inhibition for treating OAB.