Traditional Chinese Medicine For Diabetic Erectile Dysfunction: Targeting RhoA, Nrf2, MAPK, And PKC Pathways

3. RhoA/Rho Signaling Pathway

3.1 RhoA/Rho Signaling Pathway and DMED

Chitaley et al. [30] were the first to identify the expression of endogenous Rho kinase (ROCK) in penile corpus cavernosum tissue. In recent years, numerous studies have shown that the RhoA/Rho kinase pathway is involved in the development of diabetes mellitus-induced erectile dysfunction (DMED) [31–33]. RhoA in this pathway is a small GTP-binding protein belonging to the Ras-related Rho family. It primarily exists in an inactive state bound to guanosine diphosphate (GDP), acting as a molecular switch between inactive GDP-bound and active GTP-bound forms.

Rho kinase is a serine/threonine kinase with two isoforms: ROCK1 and ROCK2. These kinases can increase intracellular Ca²⁺ concentration, promoting the binding of cytoplasmic Ca²⁺ to calmodulin, thereby activating myosin light chain kinase (MLCK). MLCK then phosphorylates the myosin light chain, leading to the crosslinking of myosin and actin filaments, ultimately resulting in smooth muscle contraction and contributing to erectile dysfunction [31].

Y-27632 is a highly selective ROCK inhibitor. Studies have shown that intracavernosal injection of the ROCK inhibitor Y-27632 or an adenovirus encoding a dominant-negative mutant RhoA in aged rats can inhibit the RhoA pathway, reduce ROCK activity, and improve intracavernosal pressure both at baseline and following electrical stimulation [34]. Additionally, Y-27632 improves erectile function in castrated rats with hypogonadism and induces maximal relaxation of isolated corpus cavernosum smooth muscle tissue [35–36].

 

 

 

Herbal Cistanche Suppllement For ED

 

3.2 Traditional Chinese Medicine Intervenes in DMED via RhoA/Rho Pathway

The widespread involvement of the RhoA pathway in ED makes it a rational pharmacological target for clinical applications [37]. The Tianjing Tongluo Formula, composed of Xianlingpi, Shudihuang, Shuizhi, and Chuanniuxi, has been shown to improve endothelial function and repair vascular damage. Yu Xudong et al. [31] found that this formula may ameliorate erectile function in STZ-induced DMED rats by downregulating the expression of RhoA, ROCK1, and ROCK2 proteins in the RhoA/Rho pathway, increasing serum testosterone (T), and activating eNOS activity.

Schisandrin B, one of the lignan components in Schisandra chinensis, possesses anti-inflammatory, antioxidant, hepatoprotective, and immunoregulatory properties. Guo Pengpeng [38] observed that after 8 weeks of oral administration of schisandrin B in DMED rats, the expression of eNOS/NO-factors promoting smooth muscle relaxation in the corpus cavernosum-increased, while expression of RhoA/ROCK/phosphorylated myosin phosphatase target subunit 1 (p-MYPT-1)-factors promoting contraction-decreased. This suggests that schisandrin B may enhance erectile function by inhibiting RhoA/ROCK expression and upregulating eNOS/NO.

Buyang Huanwu Decoction, composed of Shanzhuyu, Huangqi, Taoren, Dangguiwei, Shanyao, Honghua, Chishao, Dilong, Chaihu, Dangshen, Chuanxiong, Shudihuang, Gegen, and Sanqi, has multiple effects such as promoting blood circulation, replenishing Qi, nourishing essence, and improving the penile vascular system. Clinical trials have shown that the decoction can increase serum testosterone and reduce estradiol (E2), luteinizing hormone (LH), follicle-stimulating hormone (FSH), and prolactin (PRL), thereby improving erectile function [39].

In animal studies, Buyang Huanwu Decoction downregulated the expression of RhoA, ROCK2, and transforming growth factor-beta 1 (TGF-β1) in the penile tissue of DMED rats, while upregulating alpha-smooth muscle actin (α-SMA) levels. It improved erectile function by inhibiting corpus cavernosum fibrosis and reducing inflammation [40].

These studies collectively demonstrate that active components or compound formulas such as schisandrin B, Tianjing Tongluo Formula, and Buyang Huanwu Decoction can improve DMED by inhibiting the RhoA/Rho signaling pathway. Their mechanisms include reducing oxidative stress, alleviating inflammation, preventing fibrosis in the corpus cavernosum, and relaxing smooth muscle, ultimately leading to improved erectile function.

4. Nrf2 Signaling Pathway

4.1 Nrf2 Signaling Pathway and DMED

The Nrf2/ARE signaling pathway comprises three core molecules: Nrf2, Kelch-like ECH-associated protein 1 (Keap1), and the antioxidant response element (ARE) [41]. Nrf2 is a key positive regulator of antioxidant genes in various reproductive system diseases. Its deficiency leads to a reduction in antioxidant factors and enzyme activities, resulting in an imbalance in intracellular redox homeostasis.

Keap1 is a cytoplasmic protein chaperone. Under oxidative stress, Nrf2 dissociates from Keap1 and translocates into the nucleus, where it activates the transcription of various antioxidant and phase II detoxifying enzymes to restore redox balance [42]. ARE is a specific DNA promoter binding sequence in the nucleus that can be activated by Nrf2 to regulate the expression of phase II detoxifying enzymes, antioxidant enzymes, and anti-inflammatory factors, thereby enhancing the cell's antioxidant capacity.

4.2 Intervention of DMED via Nrf2/ARE Pathway by Traditional Chinese Medicine

Dilong protein, extracted from the traditional Chinese medicine Dilong (earthworm), possesses multiple pharmacological effects, including antioxidant, anti-inflammatory, antifibrotic, anticoagulant, antihypertensive, and microcirculation-improving properties. Zhang Aiping et al. [43] found that Dilong protein could upregulate Nrf2 and SOD expression in DMED rats while downregulating MDA, NF-κB, and IL-1β protein levels, as well as NF-κB, IL-1β, and TNF-α mRNA expression. Its mechanism may involve regulation of the Nrf2/ARE signaling pathway to inhibit oxidative stress and inflammatory responses, thereby improving erectile dysfunction.

Niangmu Liangyao Wine is a traditional medicinal wine prepared by soaking Niangmu Liang-a Wa ethnic medicine from Yunnan-into white liquor. It is known for strengthening muscles and bones and tonifying Yang. Zhao Jie et al. [44] found that this wine increased the levels of nNOS, eNOS, NO, cGMP, SOD, and GSH-Px in the corpus cavernosum of rats with arteriogenic erectile dysfunction. It also upregulated both mRNA and protein expression of Nrf2 and heme oxygenase-1 (HO-1), activated Nrf2-related signaling factors, inhibited oxidative stress, and improved tissue damage and erectile function in rats.

Crocin, the main active component of Crocus sativus (saffron), has pharmacological effects such as promoting blood circulation, cooling the blood, detoxification, calming the mind, protecting the liver, and promoting bile secretion. Zang Zhenjie [45] found that crocin could upregulate the expression of α-SMA, eNOS, von Willebrand factor (vWF), Nrf2, HO-1, and Bcl-2 in the penile tissue of DMED rats, while downregulating the expression of Bax and cleaved Caspase-3. Both in vivo and in vitro experiments confirmed that crocin could alleviate oxidative stress and reduce apoptosis in the corpus cavernosum, thus improving erectile function. This effect may be related to the activation of the Nrf2/HO-1 signaling pathway.

In conclusion, the above-mentioned traditional Chinese medicine extracts and compound formulas may activate the Nrf2/ARE signaling pathway, regulate oxidative stress and downstream inflammatory responses, improve endothelial function, and increase penile microcirculation perfusion, thereby exerting therapeutic effects on DMED.

5. MAPK Signaling Pathway

5.1 MAPK Signaling Pathway and DMED

The MAPK family includes extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 kinase. The MAPK/ERK pathway is involved in cellular proliferation, differentiation, migration, senescence, and apoptosis [46]. Studies have shown that inhibiting the MAPK/ERK pathway and reversing abnormal ERK phosphorylation can protect vascular endothelial cells from diabetic damage, thereby improving erectile function [47]. Abnormal activation of this pathway may increase endothelial dysfunction, apoptosis, and fibrosis, leading to erectile dysfunction (ED) [48]. Nangle et al. [49] found that the p38 MAPK inhibitor LY2161793 could correct NO-dependent indicators of diabetic erectile autonomic neuropathy. Nunes et al. [50] reported that the ERK inhibitor PD98059 could induce relaxation of the corpus cavernosum in diabetic mice, reduce arginase activity, and reverse the diabetes-induced downregulation of eNOS, indicating that inhibition of the MAPK pathway may improve DMED.

 

5.2 Intervention of DMED via MAPK Pathway by Traditional Chinese Medicine

Berberine, an isoquinoline alkaloid extracted from traditional Chinese herbs such as Phellodendron, Coptis chinensis, and Sankezhen, exhibits antibacterial, anti-inflammatory, lipid-lowering, and hypoglycemic effects. Chen Geng et al. [51] found that berberine downregulated the expression of p-ERK and p-p38 MAPK in the corpus cavernosum of DMED rats. Its mechanism may involve inhibiting the p38 MAPK/ERK pathway, improving penile tissue fibrosis, and suppressing apoptosis.

Liu et al. [52] discovered that berberine could reduce TGF-β1 expression and fibrotic progression in high glucose-induced corpus cavernosum smooth muscle cells. Animal experiments further confirmed that berberine improved corpus cavernosum relaxation function in diabetic rats by inhibiting the ERK1/2 signaling pathway, reducing arginase activity, and increasing eNOS activity, thus suppressing apoptosis and fibrosis.

In summary, berberine may improve endothelial dysfunction, fibrosis, and apoptosis in penile tissue by inhibiting the MAPK signaling pathway. Although research on the role of traditional Chinese medicine in improving DMED via the MAPK pathway is limited, some studies have shown that JNK is involved in apoptosis of corpus cavernosum cells in rats [53], and that the ERK1/2 pathway can inhibit eNOS activity in the corpus cavernosum [54], suggesting that the MAPK signaling pathway is a potential therapeutic target for DMED.

 

This is consistent with the results observed in the PKC inhibitor LY333531 intervention group in mice, suggesting that the underlying mechanism may involve the inhibition of the PKC signaling pathway, exerting protective effects on endothelial function and antiplatelet activation, thereby alleviating erectile dysfunction. In a clinical trial involving 66 patients with diabetic erectile dysfunction (DMED), it was found that the combination of Huoxue Tongluo Qiwetang (a traditional Chinese decoction promoting blood circulation and unblocking collaterals) with a low dose of tadalafil showed higher efficacy than tadalafil alone, indicating that this decoction can enhance the therapeutic effects in DMED patients and compensate for the limitations of PDE5 inhibitors [66].

In summary, traditional Chinese medicine (TCM) shows potential in regulating the PKC signaling pathway. However, specific studies on its application in DMED remain limited. Future research should further explore the regulatory mechanisms of TCM components on the PKC pathway and their practical application in DMED treatment, aiming to provide safer and more effective therapeutic options for patients.

 

8. Conclusion and Outlook

Various TCM monomers and compound formulas can intervene in the pathogenesis of DMED through multiple signaling pathways, thereby improving symptoms, enhancing patients' quality of life, and potentially increasing fertility rates. A summary of mechanisms and therapeutic targets is provided in the supplementary materials of the extended publication.

Numerous studies have shown that:

TCM treatment for DMED does not rely on a single signaling pathway. For instance, Dilong protein can improve DMED by activating the Nrf2/ARE and NO/cGMP pathways while inhibiting the RhoA/Rho pathway; Shuizhi-Wugong Granules exert therapeutic effects by activating the PI3K/Akt and NO/cGMP pathways.

Multiple signaling pathways involved in TCM treatment of DMED often interact. For example, regulation of the PI3K/Akt pathway can activate the NO/cGMP pathway, thereby increasing NO and cGMP levels and promoting restoration of erectile function. AMPK regulates the Nrf2/HO-1 pathway, and both PI3K/Akt and AMPK pathways can modulate mTOR to influence autophagy. S1PR1 is an upstream target of both the PI3K/Akt and TGF-β/Smad pathways. Signaling molecules such as MAPK, PI3K/Akt, and PKC are widely involved in the activation and nuclear translocation of Nrf2. These pathways also regulate eNOS to exert antioxidative effects, ultimately improving DMED.

Currently, abundant research has confirmed that both TCM monomers and compound formulas can exert therapeutic effects by regulating multiple signaling pathways. These include reducing oxidative stress, inhibiting inflammation, modulating autophagy, and improving vascular endothelial dysfunction in the penis, achieving satisfactory results and demonstrating the multifaceted advantages of TCM in treating DMED.

However, several limitations remain:

Due to the varying components and dosages of different TCM herbs or formulas, the mechanisms of action may differ. Most current studies are based on empirical prescriptions with unclear active ingredients, low reproducibility, and insufficient scientific rigor.

Mechanistic studies of TCM in treating or preventing DMED often focus on surface phenomena, with insufficient in-depth exploration of signaling pathways. Most studies are based on animal models and lack large-scale clinical data, including multicenter randomized controlled trials, to verify the efficacy and safety of TCM.

Most animal intervention studies last only 4–8 weeks, and the absorption and metabolism of TCM in animals may not be ideal.

TCM treatment typically emphasizes holistic regulation and syndrome differentiation, which may reduce efficacy in patients needing rapid symptom relief due to poor compliance or individual variation.

Future research should focus on exploring the optimal ratios of various TCM formulations, combination therapies with other treatment modalities, and conducting high-quality clinical trials to develop personalized and comprehensive treatment plans for DMED patients.

 

In Summary

TCM demonstrates multi-pathway, multi-target, clinically effective, and highly safe characteristics in the treatment of DMED. It presents a broad application prospect in the intervention of diabetic erectile dysfunction. With ongoing research and technological advancements, TCM is expected to bring new hope to more DMED patients and improve their quality of life.