Electroacupuncture improves cyclophosphamide-induced bladder overactivity by reducing mechanotransduction in the rat urothelium

doi:10.19852/j.cnki.jtcm.2025.02.003

Abstract

Abstract:

OBJECTIVE: To investigate whether electroacupuncture (EA) therapy for overactive bladder (OAB) exerts its effect by modulating mechanosensitive channels in the urothelium, thereby improving bladder sensory function.

METHODS: In this study, a rat model of OAB was created by using intraperitoneal injections of cyclophosphamide. We performed either EA or bladder perfusion with HC-067047 [a transient receptor potential vanilloid 4 (TRPV4) antagonist] and assessed the efficacy of electro-acupuncture in the treatment of OAB in rats viaurodynamic determination and Void spot assay. tissue morphology, distribution and expression of the TRPV4 protein and the amount of adenosine triphosphate (ATP) and Ca2+ released from urothelial cells in each group of rats were observed to identify the mechanism by which electroacupuncture improves OAB in rats.

RESULTS: EA ameliorated bladder function and voiding behaviour, improved bladder uroepithelial tissue morphology, and significantly reduced the immun-ofluorescence intensity and the mRNA and protein expression levels of TRPV4 in the uroepithelium of OAB rats. Moreover, the simulated mechanical stimulation-induced increases in Ca²⁺ concentration and the release of ATP and acetylcholine (Ach) from bladder urothelial cells were inhibited. The changes in EA followed the same trend as those in HC-067047.

CONCLUSIONS: These results suggest that EA inhibits bladder sensory function by downregulating the expression of mechanically activated TRPV4 ion channels distributed in bladder urothelial cells, which correspondingly decreases the inward flow of extracellular Ca²⁺ and reduces the release of ATP and Ach, thereby attenuating excitatory signals.

Key words: electroacupuncture, urinary bladder, overactive, urothelium, TRPV cation channels, mechanotransduction, cellular

HU Junwei, FENG Jiwei, LI Wen, LIU Lumin, LI Xu, XU Ge, LIU Jiandang, CHEN Yuelai. Electroacupuncture improves cyclophosphamide-induced bladder overactivity by reducing mechanotransduction in the rat urothelium[J]. Journal of Traditional Chinese Medicine, 2025, 45(2): 348-358.

Figures/Tables 8

Table 1 Comparison of urodynamic evaluation ($\bar{x}±s$)

Group	n	Storage time (min)			Bladder capacity (mL)		Voiding pressure (cmH₂O)
Group	n	Model establishment	After intervention		Model establishment	After intervention	Model establishment	After intervention
Control	10	6.02±0.68		6.11±0.93	1.51±0.17	1.53±0.23	58.50±6.87	59.60±7.24
Model	10	3.07±1.03^a		2.63±0.74^a	0.77±0.26^a	0.68±0.19^a	46.20±6.56^a	43.60±4.79^a
EA	10	3.06±1.22^a		5.38±1.12^bc	0.77±0.31^a	1.35±0.28^bc	47.00±4.16^a	52.60±4.27^abc
HC-067047	10	3.20±0.87^a		5.74±1.11^bc	0.80±0.22^a	1.43±0.28^bc	46.90±7.03^a	53.40±4.99^abc

Table 2 Comparison of voiding behavior ($\bar{x}±s$)

Group	n	The number of urinations in 3 h	Total urinary output in 24 h
Control	10	7.3±1.5	20.0±1.9
Model	10	18.9± 2.2^a	7.9±1.8^a
EA	10	10.0±1.2^ab	15.1±1.6^ab
HC-067047	10	9.5±1.1^ab	16.0±1.7^ab

Table 3 Comparison of bladder weight and the expression of TRPV4 in urothelium ($\bar{x}±s$)

Group	n	Bladder weight (mg)	Fluorescent intensity (AU)	Protein (TRPV4/β-Actin)	mRNA (TRPV4/β-Actin)
Control	10	86.0246±2.7341	0.0419±0.0034	0.3046±0.0451	3.0233±0.7845
Model	10	110.5827±3.7873^a	0.0517±0.0043^a	0.8840±0.0694^a	7.7579±1.7223^a
EA	10	93.1403±4.2128^ab	0.0461±0.0046^b	0.6490±0.0408^ab	4.1362±0.9476^b
HC-067047	10	93.9763±2.6271^ab	0.0441±0.0028^b	0.4303±0.0270^ab	3.3301±0.4104^b

Figure 1 Effect of EA on OAB symptoms in rats A1-A4: representative urine spot plots. B1-B4: urodynamic diagram. A1, B1: no modeling or treatment was administered. A2, B2: a single intraperitoneal injection of 2.5% cyclophosphamide (150 mg/kg) without any treatment. A3, B3: EA: treatment once daily for three consecutive days after molding. A4, B4: HC-067047 (TRPV4 antagonist) (0.2 mL/time) was instilled into the bladder for 3 consecutive days after molding. EA: electroacupuncture; OAB: overactive bladder; TRPV4: transient receptor potential vanilloid 4.

Figure 2 Bladder urothelium tissue structure [HE staining of the bladder wall (×20)] A: control; B: Model; C: EA; D: HC-067047. Control: no modeling or treatment was administered; Model: a single intraperitoneal injection of 2.5% cyclophosphamide (150 mg/kg) without any treatment. EA: EA treatment once daily for three consecutive days after molding. HC-067047 (TRPV4 antagonist): HC-067047 (0.2 mL/time) was instilled into the bladder for 3 consecutive days after molding. EA: electroacupuncture; HE staining: hematoxylin and eosin staining; TRPV4: transient receptor potential vanilloid 4.

Figure 3 TRPV4 expression in the bladder urothelium A1-A4 (× 20): DAPI dyeing; B1-B4 (× 20): immunofluorescence assay: Alexa Fluor ?. Cy-7 dyeing; C1-C4 (× 20): immunofluorescence assay: Alexa Fluor ?. TRPV4 dyeing; D1-D4 (× 20): merge of DAPI dyeing, Cy-7 dyeing and TRPV4 dyeing. A1, B1, C1, D1: control; A2, B2, C2, D2: Model; A3, B3, C3, D3: EA; A4, B4, C4, D4: HC-067047. Control: no modeling or treatment was administered; Model: a single intraperitoneal injection of 2.5% cyclophosphamide (150 mg/kg) without any treatment. EA: EA treatment once daily for three consecutive days after molding. HC-067047 (TRPV4 antagonist): HC-067047 (0.2 mL/time) was instilled into the bladder for 3 consecutive days after molding. TRPV4: transient receptor potential vanilloid 4; DAPI: 4',6-diamidino-2-phenylindole; EA: electroacupuncture.

Figure 4 TRPV4 protein expression in the bladder urothelium Control: no modeling or treatment was administered; model: a single intraperitoneal injection of 2.5% cyclophosphamide (150 mg/kg) without any treatment. EA: EA treatment once daily for three consecutive days after molding. HC-067047 (TRPV4 antagonist): HC-067047 (0.2 mL/time) was instilled into the bladder for 3 consecutive days after molding. TRPV4: transient receptor potential vanilloid 4; EA: electroacupuncture.

Table 4 Comparison of the mechanical sensitivity of urothelial cells ($\bar{x}±s$)

Group	n	Ca²⁺(F1/F0)	The release of ATP (nM)			The release of Ach (nM)
Group	n	Ca²⁺(F1/F0)	Isotonic state	Hypotonicity state	F1/F0	Isotonic state	Hypotonicity state	F1/F0
Control	10	0.83±0.44	2.20±0.50	4.49±1.19^c	1.08±0.54	10.57±2.37	20.88±5.82^c	0.95±0.13
Model	10	2.99±0.92^a	4.66±1.03^a	18.38±4.40^ac	3.13±1.32^a	19.81±3.28^a	79.10±9.16^ac	2.94±0.35^a
EA	10	1.52±0.75^ab	3.94±1.05^ab	10.68±1.73^abc	1.88±0.80^ab	14.69±4.15^ab	44.55±8.15^abc	2.12±0.44^ab
HC-067047	10	1.45±0.66^ab	3.72±1.28^ab	10.08±2.80^abc	1.85±0.72^ab	14.51±3.53^ab	43.35±8.76^abc	2.03±0.31^ab

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