NF-κB, which interacts closely with inhibitory proteins (IκB) and IκB kinase (IKK), is a key intracellular molecule that mediates the inflammatory response. IκB and IKK are two essential upstream modulatory elements of the NF-κB signal transduction cascade. IKK-β is a protein subunit of IκB-α that can bind to cytoplasmic NF-κB and keep it inactive. After translocating into the nucleus, NF-κB is activated and promotes the transcription of proinflammatory factors such as TNF-α, TNF-α, IL-1β, IL-6 and MCP-1 (Hong et al., 2017).

In a rat model of IgA nephropathy induced by bovine serum albumin (BSA) gavage, LPS, and CC14 injection, the oral administration of AR (3 g/kg/d for 6 weeks) was found to significantly reduce urinary erythrocyte count, urinary protein, and β-N-acetylaminoglucosidase (NAG) content, and also alleviate renal tubulointerstitial lesions in IgAN rats. These beneficial effects are thought to be attributed to the downregulation of NF-κB and MCP-1 expression (Zhang et al., 2008). Moreover, AR injection (2, 20, 200 μg/L for 48 h, 72 h) was shown to inhibit high glucose-induced apoptosis and NF-κB protein expression in renal tubular epithelial cells (HK-2). The inhibitory effect was found to be positively correlated with the dose. Further investigations revealed that intraperitoneal injection of AR injection (1 mL/week for 7 weeks) reduced the expression rate of NF-κB and cytokine TNF in rats with immune complex nephritis model established by tail vein injection with BSA. This suggests that AR injection may exert therapeutic effects by suppressing the immune response through the reduction of NF-κB expression (Wang et al., 2001; Mei et al., 2016). Additionally, in the lipopolysaccharide (LPS)-induced diabetic nephropathy (DN) rat model, AR decoction (1.5, 3.0 g/kg for 8 weeks) was found to significantly improve general indices, renal function, and proteinuria. It was also observed to decrease the phosphorylation of IKK and NF-κB by reducing the expression of TLR4, thus reducing downstream inflammatory factors such as IL-6, TNF-α, and IL-1β expression levels. These results suggest that AR decoction may exert an inhibitory effect on TLR4/NF-κB signaling pathway, thereby reducing the inflammatory response and improving renal pathological indices (Li, 2019).

TLRs are a family of germline-encoded receptors that are responsible for the development of inflammatory and immune responses. TLRs are expressed on various cells, including antigen-presenting cells and intrinsic kidney cells. The recognition of TLR ligands prompts the innate immune response and stimulates TLR signaling, which initiates M1 macrophage polarization and infiltration and mediates the transcription of NF-κB, which is followed by the inflammatory cascade and proinflammatory cytokine and chemokine release. Notably, almost all TLRs (except for TLR3) use myeloid differentiation primary response 88 (MYD88) as a general adapter protein to activate NF-κB. Activation of the TLR signaling pathway has been reported to exacerbate inflammation (Zhang T. et al., 2017; Zhao and Han, 2018).

Administration of SHT (1.5, 3.0 g/kg for 7 days prior to pre-operation) significantly decreased the protein expression and RNA levels of TLR2 and TLR4 in rats with renal ischemia-reperfusion injury (IRI), and its improvement of AKI symptoms may be achieved by reducing the expression of these renal immune inflammatory receptors (Zheng et al., 2012). The effects of SHT on inflammatory cytokines associated with TLR2 and TLR4, as well as MyD88, were further investigated. The results showed that SHT intervention significantly reduced the levels of IL-6, IL-12, and MyD88 protein and mRNA levels in the kidneys of rats, and also led to a decrease in serum levels of IL-8 and INF-γ (Li et al., 2019). In another study, AS-IV (20, 40 mg/kg for 1 week) was found to significantly reduce fibronectin (FN) and collagen I (COL-1) levels, as well as inhibit inflammatory cell infiltration and cytokine secretion in a mouse model of UUO-induced renal fibrosis. In vitro experiments with human proximal tubular epithelial cells (HK-2 cells) stimulated with LPS (100 ng/mL) also demonstrated that AS-IV (10, 20 μM for 48 h) inhibited TNF-α and IL-1β expression, and these results suggest that AS-IV may protect against the progression of renal fibrosis by suppressing inflammation through the TLR4/NF-κB signaling pathway. Furthermore, AS-IV was found to decrease NF-κB expression and increase IκBα expression (Zhou et al., 2017).

The PI3K/AKT signaling pathway is vital for cell proliferation, growth and viability. As an upstream modulator of NF-κB, PI3K/AKT signaling pathway activation has been demonstrated to ameliorate inflammation (Hong et al., 2017).

Treatment with AR (5 g/kg for 8 weeks) resulted in a reduction of 24-hour urinary protein levels in DN rats, with its efficacy becoming progressively evident as the duration of treatment was extended. The underlying mechanism may involve the reduction of expression levels of p-PI3K, p-Akt, NF-κB, MCP-1, and mRNA to regulate the PI3K/Akt/NF-κB signaling pathway in DN rats, thereby attenuating renal injury (Shen, 2016). In a similar vein, treatment with AS-IV (20, 40, 80 mg/kg for 8 weeks) in a rat model of type 2 diabetes, established through intraperitoneal injection of streptozotocin (STZ) (35 mg/kg), resulted in improvements in body mass, renal index, blood glucose, 24-hour urine protein, urine microalbumin, urine creatinine, blood creatinine, and blood urea nitrogen levels when compared to the model group. Moreover, renal tissue lesions showed significant improvements, and medium and high doses of AS-IV inhibited expression of p-PI3K/PI3K, p-Akt/Akt, and p-FoxO1/FoxO1. AS-IV also upregulated the expression of BNIP3, LC3-II/LC3-I, and Beclin1, which may offer new perspectives for the clinical treatment of diabetic nephropathy by inhibiting the PI3K/Akt pathway (Ma et al., 2019).

The Wnt signaling pathway is controlled by Wnt, which is highly conserved in cells and is involved in many biological processes. To maintain the balance of transduction pathways, Wnt can act through the signaling cascade, and control cell differentiation, migration, apoptosis, and cancerous changes to control the growth and development, disease, and death (Long et al., 2010). Wnt proteins maintain a state of dynamic balance. Dysregulated expression of signaling pathway proteins can lead to various diseases, such as inflammation, carcinogenesis, neurological diseases, respiratory diseases, and metabolic disorders (Koziński and Dobrzyń, 2013).

Wang et al. discovered that overexpression of miR-21 causes the activation of protein expression of β-catenin, which, in turn, promotes renal cell fibrosis through the TGF-β1/Smad pathway in diabetic mice. AS-IV has a therapeutic effect because it downregulates the expression of miR-21 in renal cells. However, the therapeutic effect of AS-IV is suppressed when either the Wnt/β-catenin pathway or TGF-β1/Smad pathway is inhibited. Thus, AS-IV is considered a promising drug for the treatment of glomerular diseases (Wang et al., 2018). Furthermore, Li et al. demonstrated that the Wnt/β-catenin signaling pathway was abnormally activated in the kidneys of UUO rats. AS-IV (3.3, 10, 33 mg/kg for 14 days prior to the operation) inhibited this effect in a concentration-dependent manner and suppressed renal interstitial fibrosis (Wang et al., 2014a). Other studies have also found that AS-IV and APS can decrease the expression of inflammatory factors, such as IL-6, IL-8, and TNF-α, oxidative factors like ROS and MDA, and the apoptotic factor caspase-3 in inflammatory cells. Furthermore, the expression of β-catenin is activated in inflammatory cells for cytoprotection (Fan, 2021). AR treatment (2.1 g/kg for 12 weeks) in STZ-induced DN rats can reduce 24-hour urine protein quantification, attenuate interstitial pathological damage, and inhibit the expression of wnt4, β-catenin, and TGF-β1 in the renal interstitium, thus playing a role in kidney protection and slowing down the process of interstitial fibrosis in DN rats (Deng and Fang, 2012).

The JAK-STAT signaling pathway comprises three primary components, including tyrosine kinase-related receptors, the tyrosine kinase JAK, and the transcription factor STAT (Banerjee et al., 2017). In mammals, the JAK family constitutes a non-receptor class of tyrosine kinases that consists of JAK1, JAK2, JAK3, and TYK2. Among these, JAK1, JAK2, and TYK2 are broadly expressed in various tissues and cells, while JAK3 is restricted to bone marrow cells and lymphocytes (Chuang and He, 2010). STATs are a group of cytoplasmic proteins that interact with the regulatory regions of target genes, located downstream of JAKs, and crucially participate in signal transduction and transcriptional activation. The STAT proteins comprise STAT1, STAT2, STAT3, STAT4, STAT5a, STAT5b, and STAT6. JAK-STAT signal transduction and transcriptional activation are the primary regulatory pathways associated with cytokines. Specifically, JAK binds to the intracellular domain of the activated cytokine receptor and transmits the signal to the promoter of the target gene upon activation (Xin et al., 2019).

The immune complex deposition-induced activation of the inflammatory response cascade is a major contributor to kidney injury, with inflammatory cytokines playing a crucial role in the development of chronic nephritis. SHT contains components with immune-enhancing and anti-inflammatory properties, and intervention with SHT (45 g/kg for 4 weeks) was found to decrease the expression and activation levels of STAT3 in the kidney tissue of rats with chronic serum disease nephritis, consequently reducing downstream TIMP-1 expression and inhibiting thylakoid cell proliferation. These findings suggest that SHT have a potential to alleviate renal injury in rats with nephritis (Zhang et al., 2005). Furthermore, in high glucose-treated renal tubular epithelial cells, APS (200 mg/L for 48 h) was shown to effectively inhibit apoptosis, transdifferentiation, and reactive oxygen levels. Further analysis revealed that APS treatment resulted in the downregulation of E-cadherin, α-SMA, p-STAT1, and p-STAT3 protein expressions in renal tubular epithelial cells, indicating that APS may exert its protective effect on renal tubular epithelial cells by regulating the JAK-STAT signaling pathway (Guo et al., 2018).

Innate immune cells are a vital constituent of the innate immune system, which promptly reacts to invading pathogens either after or concurrently with the fortification of tissue barriers. These cells generate non-specific anti-infective immunity, consisting of neutrophils, macrophages, and natural killer (NK) cells.

Adaptive immunity encompasses a complex system of immune organs and specialized immune cells, representing the final and most rigorous line of immune defense.

Glomerular mesangial cells, which are intrinsic cells located in the glomerulus between capillary loops, are adjacent to endothelial cells or the basement membrane and exhibit an irregular shape. Their cell processes can be deeply located between endothelial cells and the basement membrane or can extend through endothelial cells into the capillary lumen. Glomerular mesangial cells perform various functions such as the secretion of extracellular matrix, production of cytokines, support of the glomerular capillary plexus, phagocytosis and removal of macromolecular substances, and contraction similar to smooth muscle cells.

Interstitial proliferative nephritis is a prevalent form of chronic glomerulonephritis. Research has indicated that after a duration of 3 months of administering SHT (0.75, 1.5, 3 g/kg), the levels of urine protein, serum creatinine, and total cholesterol showed a significant decrease. Furthermore, serum total protein showed an increase, while the number of PCNA-positive cells in interstitial cells decreased significantly in rats. These findings indicate that SHT is effective in ameliorating the symptoms of Thy1 nephritis and inhibiting the proliferation of interstitial cells in rats (Geng et al., 2016). Additionally, Wang et al. discovered that AS-IV increased the expression of CHOP, BAX, and cleaved caspase3, which regulate apoptosis, by upregulating the phosphorylation of endoplasmic reticulum stress-related proteins eIF2α, PERK, and IRE1α, thereby effectively alleviating clathrin-induced apoptosis in rat thylakoid cells (Wang et al., 2022). When AS-IV (50 μg/mL for 72 h) was administered to RMC cultured in high glucose, it downregulated miR-192 expression and inhibited high glucose-induced RMC overproliferation. In a rat DN model induced by STZ, AS-IV (40 mg/kg, for 8 weeks) also downregulated miR-192 expression and reduced the mRNA and protein levels of TGF-β1, Smad3, α-SMA, and COL-1, while increasing the mRNA and protein expression of Smad7. Therefore, AS-IV may treat DN by inhibiting interstitial hyperplasia and renal fibrosis through the TGF-β1/Smad/Mir-192 signaling pathway (Mao et al., 2019).

Podocytes represent a specialized subtype of differentiated epithelial cells that are responsible for upholding the structural and functional integrity of the glomerular filtration barrier, and furthermore determine the size of filtration proteins. When podocytes undergo apoptosis and are consequently lost, the filtration barrier structure collapses, which in turn leads to proteinuria (Lin et al., 2020; Wang et al., 2020).

The nephrotic syndrome refers to a urinary protein production caused by damage to podocytes, whereby the podocyte septum gap (known as the slit diaphragm or SD) and skeletal structure serve as essential components for maintaining filtration function. Following SHT (2.5, 5 and 10 g/kg for 12 weeks) intervention in a rat model of adriamycin nephropathy, the protein expression and RNA levels of SD-related proteins, namely, podocin and nephrin, were significantly upregulated in kidney tissues. Additionally, the protein expression and RNA levels of the podocyte skeleton-related protein CD2AP and the cytoskeletal damage component desmin were significantly decreased. These findings suggest that SHT exerts its mechanism of action on renal protection in rats with adriamycin nephropathy by improving glomerular podocyte lytic septum protein, maintaining podocyte skeleton homeostasis, and promoting glomerular podocyte repair after injury (Wang, 2016). AS-IV (6, 8, and 18 mg/kg for 2 weeks) improved renal function in db/db mice, reduced podocyte injury, and increased glomerular podocyte Klotho expression in glomerular foot cells (Xing et al., 2021). AS-II (at doses of 3.2 and 6.4 mg/kg for 9 weeks) improved STZ-induced proteinuria, renal histopathology, foot cell degeneration, and foot cell apoptosis in diabetic rats. This treatment partially restored the expression of renal mitochondrial dynamics-related and autophagy-related proteins, including Mfn2, Fis1, P62, and LC3. Moreover, AS-II increased the expression of PINK1 and Parkin, which are associated with mitochondrial autophagy, suggesting that AS-II may improve DN by regulating the Nrf2 and PINK1 pathways in diabetic rats (Gao et al., 2020; Su et al., 2021). AR was found to inhibit the expression of TGF-β1, FN, and MDA and increase SOD activity, TRPS expression, and the levels of α-dystroglycan and integrin to protect podocytes from damage (Wu and Li, 2016). Furthermore, AS-II downregulated TGF-β1, Smad3, and beta-catenin by inhibiting the expression of miR-21 and upregulating Smad7 expression to attenuate podocyte dedifferentiation (Cong, 2021). Additionally, AS-IV upregulated lncRNA-TUG1 and downregulated TRAF5 expression to reduce high glucose-induced podocyte injury (Lei et al., 2018). Xiao et al. investigated the effects of AS-IV on mouse podocytes and found that AS-IV promoted the connection between podocyte peduncles, improved their structural disorder, and alleviated glomerular basement membrane (GBM) thickening, while inhibiting excessive ECM. AS-IV downregulated TGF-β, N-cadherin, and α-SMA levels in podocytes and upregulated nephrin, E-cadherin levels, and SIRT1 expression. Furthermore, AS-IV reduced p65 acetylation levels and alleviated high glucose-induced podocyte EMT12 by enhancing autophagy, with the mechanism potentially acting through the regulation of the SIRT1-NF-κB pathway (Wang et al., 2019).

Glomerular endothelial cells are intrinsic cells within the glomerulus and a crucial constituent of the nephron. They have the capacity to modulate glomerular filtration, playing a crucial role in the inner lining of the filtration barrier and are intricately involved in the pathophysiological processes of the glomeruli.

Administration of AR injection has been demonstrated to effectively prevent and repair STZ-induced vascular endothelial cell tight junction damage in diabetic rats, as well as reduce such injury in rats induced by high glucose peritoneal dialysis fluid (Shu et al., 2007). AR has been shown to decrease the release of endothelin, improve inflammation and oxidative stress in endothelial cells, balance intraglobular coagulation, and prevent apoptosis. This is accomplished through inhibiting the TNF-α/NF-κB signaling pathway, upregulating stem cell regulatory factors such as Nanog and Sox2, and inhibiting the expression of CAM, ROS, Fas, and MDA (Zhang and Que, 2020). Moreover, AR mono-AR injection has been found to upregulate the expression of sFlt-1 and downregulate VEGF, thereby promoting the proliferation and migration of glomerular endothelial cells (Cen et al., 2016). After treatment with AR combination (20 mL/kg for 12 weeks), unilateral nephrectomized and STZ-induced DN rats exhibited different biological indices in the glomeruli, including urine volume, blood glucose, glycosylated hemoglobin, lipids, VACM-1 and α-SMA. This treatment may protect renal endothelial cells by inhibiting VCAM-1 expression and preventing endothelial cell phenotypic transformation, thereby safeguarding renal tissue in DN rats (Zhang et al., 2014).

Renal tubular epithelial cells represent the most fundamental structural units of renal tubules and play an essential role in the physiological function of the kidneys. Renal tubules are divided into proximal, distal, and fine segments, each with a unique epithelial cell function. Proximal renal tubular epithelial cells are responsible for reabsorbing glucose, amino acids, and proteins filtered from the glomerulus, as well as secreting carbonate ions to regulate the acid-base balance in the body. Distal renal tubular epithelial cells, on the other hand, mediate processes such as dilution, acidification, and concentration, resulting in a decrease in the specific gravity of urine. These cells also secrete hydrogen ions to regulate the acid-base balance in the body, thus maintaining homeostasis. Finally, the fine epithelial cells are the thinnest part of the renal tubule, and they are responsible for reabsorbing water, sodium, and urea. These cells are located between the proximal and distal tubules, are divided into descending and ascending branches, and can participate in the formation of the nephron medullary loop.

Injury to renal tubular epithelial cells is a major pathological process during ischemia-reperfusion injury (IRI) of the kidney. The localization and distribution of the sodium-potassium pump (Na⁺-K⁺-ATPase) within the renal tubular epithelium is a crucial factor affecting renal tubular function. In IRI, disruption of the Na⁺-K⁺-ATPase skeleton leads to a loss of epithelial cell function. SHT (1.5, 3.0 g/kg for 7 days) intervention after IRI resulted in an observed upregulation of Na⁺-K⁺-ATPase expression levels and improved distribution (Wei et al., 2013). Additionally, the sodium/dicarboxylic acid co-transporter protein (NaDC1), located primarily on the brush border side of the tubule, is a tubular polarity marker that acts in synergy with Na⁺-K⁺-ATPase to aid in the absorption of tricarboxylic acid cycle intermediates. Studies indicate that NaDC1 expression is decreased in renal tubular epithelial cells during IRI, resulting in a loss of tubular reabsorption function. A previous study indicated that intervention in a rat tubular epithelial cell hypoxia/reoxygenation model using serum containing SHT drug resulted in improved intracellular NaDC1 expression and distribution, suggesting that SHT could promote tubular reabsorption and repair tubular injury (Yang et al., 2014). AS-IV effectively inhibited UUO and TGF-β1-induced apoptosis in renal tubular epithelial cells by inhibiting Caspase-3 activation to prevent injury (Wang et al., 2014b). The anti-fibrotic mechanism of AS-IV prevents renal tubular epithelial cell apoptosis primarily through inhibition of Caspase-3 activation, and it may be mediated by downregulation of phosphorylated p38 and phosphorylated c-JunN-terminal kinase (Xu et al., 2014). Furthermore, APS inhibits the expression of MMP-2 and downregulates the expression of TIMP-1, TNF-α, and AngII (Lu and Wei, 2014; Sun et al., 2021). In addition, AS-IV inhibits multiple signaling pathways, including P38MAPK (Chen and Xie, 2019), TGF-β1/1-ILK (Zheng et al., 2021), and JAK/STAT (Guo et al., 2018). It also reduces the phosphorylation of P65 protein and effectively inhibits immune inflammation, apoptosis, and degradation of the extracellular matrix (ECM) in the renal tubular epithelium.