Bushen Kaixuan Tongluo Formula Ameliorates Diabetic Kidney Disease in Db/db Mice Via Activation Of The CAMP/PKA/CREB–Bcl-2 Pathway And Improvement Of Glucose–Lipid Metabolism
Oct 15, 2025
3 Results
3.1 Effects of Bushen Kaixuan Tongluo Formula on general conditions of db/db mice
Mice in the blank group were active with glossy fur. Mice in the model group were obese, listless, and showed markedly reduced activity time. Mice in all medication groups showed certain improvements compared with the model group.
TCM HERB CISTANCHE EXTRACT FOR KIDNEY HEALTH
3.2 Effects of Bushen Kaixuan Tongluo Formula on body weight of db/db mice
During the experimental period, compared with the blank group, body weight in the model group increased significantly (P<0.01). Compared with the model group, body weight in all medication groups tended to decrease, with differences appearing from week 8; the high-dose Bushen Kaixuan Tongluo Formula group showed the best effect. See Table 2.
Table 2. Effect of Cistanche-Based Formula on Body Weight in db/db Mice (x̄±s, n = 6)
| Group | Dose (g·kg⁻¹) | Week 0 (g) | Week 4 (g) | Week 8 (g) | Week 12 (g) |
|---|---|---|---|---|---|
| Blank | - | 24.38 ± 0.35 | 29.88 ± 0.46 | 32.92 ± 0.90 | 33.14 ± 0.49 |
| Model | - | 37.27 ± 1.46 | 45.63 ± 1.85¹ | 49.38 ± 3.52² | 50.86 ± 4.69² |
| Low-dose formula | 7 | 36.47 ± 1.40 | 44.07 ± 1.84 | 43.41 ± 3.11 | 40.03 ± 1.36³ |
| Medium-dose formula | 14 | 37.57 ± 1.01 | 44.23 ± 2.05 | 42.47 ± 3.36³ | 40.25 ± 4.78³ |
| High-dose formula | 28 | 38.03 ± 1.62 | 43.28 ± 1.63 | 40.82 ± 1.87⁴ | 36.49 ± 1.32⁴ |
| Irbesartan | 0.02 | 38.58 ± 1.66 | 44.63 ± 1.66 | 46.03 ± 2.10 | 44.83 ± 2.43 |
3.3 Effects of Bushen Kaixuan Tongluo Formula on renal function of db/db mice
Compared with the blank group, Scr in the model group showed an upward trend without statistical significance, while BUN, uALB, and ACR levels were significantly elevated (P<0.05). Compared with the model group, Scr in the formula groups tended to decrease without statistical significance; BUN, uALB, and ACR were significantly reduced (P<0.05), with the medium-dose formula group showing the best effect. In the irbesartan group, Scr and BUN tended to decrease without statistical significance, while uALB and ACR decreased significantly (P<0.01). See Table 3.
Table 3. Effect of Cistanche-Based Formula on Renal Function in db/db Mice (x̄±s, n = 6)
| Group | Dose (g·kg⁻¹) | Scr (μmol/L) | BUN (mmol/L) | uALB (mg/L) | ACR (mg/g) |
|---|---|---|---|---|---|
| Blank | - | 53.76 ± 0.91 | 6.79 ± 0.26 | 1.69 ± 0.13 | 9.54 ± 1.01 |
| Model | - | 61.34 ± 1.89 | 9.08 ± 0.73¹ | 4.16 ± 0.19² | 63.08 ± 9.92² |
| Low-dose formula | 7 | 55.69 ± 3.55 | 6.99 ± 0.54³ | 1.81 ± 0.05⁴ | 21.37 ± 0.62⁴ |
| Medium-dose formula | 14 | 50.45 ± 3.02 | 6.48 ± 0.26⁴ | 1.21 ± 0.08⁴ | 19.99 ± 2.03⁴ |
| High-dose formula | 28 | 51.59 ± 3.09 | 6.01 ± 0.30⁴ | 1.34 ± 0.09⁴ | 22.72 ± 1.38⁴ |
| Irbesartan | 0.02 | 55.33 ± 2.31 | 7.93 ± 0.48 | 1.37 ± 0.09⁴ | 27.00 ± 3.83⁴ |
3.4 Effects of Bushen Kaixuan Tongluo Formula on liver function and lipid metabolism of db/db mice
Compared with the blank group, ALT, AST, TC, and TG levels in the model group increased significantly (P<0.05). Compared with the model group, AST and ALT in the formula groups showed downward trends without statistical significance, TC decreased significantly (P<0.01), TG in the medium-dose formula group decreased significantly (P<0.05), and other TCM groups showed a downward trend in TG without statistical significance. In the irbesartan group, TC showed a downward trend without statistical significance, while TG decreased significantly (P<0.01). See Table 4.
Table 4. Effect of Cistanche-Based Formula on Liver Function and Lipid Metabolism (x̄±s, n = 6)
| Group | Dose (g·kg⁻¹) | AST (U/L) | ALT (U/L) | TC (mmol/L) | TG (mmol/L) |
|---|---|---|---|---|---|
| Blank | - | 158.80 ± 6.44 | 56.31 ± 3.00 | 2.40 ± 0.08 | 1.22 ± 0.11 |
| Model | - | 310.20 ± 53.31¹ | 206.70 ± 24.98² | 2.98 ± 0.21¹ | 3.71 ± 0.22² |
| Low-dose formula | 7 | 202.50 ± 22.27 | 125.30 ± 8.74 | 2.09 ± 0.06⁴ | 3.13 ± 0.18 |
| Medium-dose formula | 14 | 263.10 ± 31.37 | 166.80 ± 34.28 | 1.94 ± 0.10⁴ | 2.23 ± 0.32⁴ |
| High-dose formula | 28 | 266.20 ± 22.96 | 166.80 ± 22.61 | 1.97 ± 0.05⁴ | 2.96 ± 0.26 |
| Irbesartan | 0.02 | 215.30 ± 14.43 | 101.70 ± 20.41² | 2.50 ± 0.19 | 2.42 ± 0.31⁴ |
3.5 Effects of Bushen Kaixuan Tongluo Formula on glucose metabolism of db/db mice
Compared with the blank group, levels of FBG, LEP, GSP, and INS were significantly increased in the model group (P<0.05). Compared with the model group, FBG in the medication groups showed no downward trend; LEP in the formula groups tended to decrease without statistical significance, while LEP in the irbesartan group decreased significantly (P<0.05). GSP and INS decreased significantly in all medication groups (P<0.05), with the high-dose formula group showing the best effect. See Table 5.
Table 5. Effect of Cistanche-Based Formula on Glucose Metabolism in db/db Mice (x̄±s, n = 6)
| Group | Dose (g·kg⁻¹) | FBG (mmol/L) | LEP (ng/mL) | GSP (μmol/L) | INS (μIU/mL) |
|---|---|---|---|---|---|
| Blank | - | 4.17 ± 0.26 | 5.32 ± 0.18 | 127.30 ± 2.69 | 7.99 ± 0.13 |
| Model | - | 18.87 ± 3.84² | 6.46 ± 0.22¹ | 239.40 ± 14.98² | 23.34 ± 2.16² |
| Low-dose formula | 7 | 28.22 ± 2.30 | 5.60 ± 0.30 | 201.80 ± 4.68⁴ | 18.52 ± 0.72³ |
| Medium-dose formula | 14 | 28.00 ± 3.33 | 5.98 ± 0.46 | 193.30 ± 5.79⁴ | 15.64 ± 0.33⁴ |
| High-dose formula | 28 | 33.30 ± 0.00⁴ | 5.50 ± 0.08 | 178.70 ± 2.63⁴ | 14.57 ± 0.23⁴ |
| Irbesartan | 0.02 | 25.62 ± 2.36 | 5.27 ± 0.14³ | 163.80 ± 2.32⁴ | 14.28 ± 0.57⁴ |
3.6 Effects of Bushen Kaixuan Tongluo Formula on renal histopathology of db/db mice
HE staining showed normal renal structure in the blank group: regular glomeruli and intact tubular epithelial cells. The model group exhibited irregular glomerular morphology, moderate mesangial cell proliferation, inflammatory cell infiltration, and vacuolar degeneration of tubular epithelial cells. These pathological changes were alleviated to varying degrees in the low-, medium-, and high-dose formula groups and the irbesartan group. (Image converted to text summary.)
PAS staining showed a small amount of extracellular matrix (ECM) deposition in glomeruli of the blank group, while the model group exhibited massive ECM deposition and markedly thickened basement membranes. These pathological features were improved to some extent in all three dose groups of the formula and in the irbesartan group. (Image converted to text summary.)
Masson staining showed normal glomerular structure with minimal pathological collagen deposition in the blank group. The model group displayed extensive blue collagen fiber deposition in glomeruli and interstitium. The formula dose groups and the irbesartan group showed improvements of varying degrees. (Image converted to text summary.)
3.7 Effects of Bushen Kaixuan Tongluo Formula on p-PKA and p-CREB expression in renal tissue of db/db mice
Compared with the blank group, staining intensities of p-PKA and p-CREB in renal tissue of the model group were significantly reduced (P<0.01). Compared with the model group, p-PKA expression in the medium-dose formula group increased significantly (P<0.01), and p-CREB showed an upward trend without statistical significance. See Figure 4 and Table 6. (Image converted to text summary.)
3.8 Effects of Bushen Kaixuan Tongluo Formula on protein expression of p-PKA, PKA, p-CREB, CREB, and Bcl-2 in renal tissue of db/db mice
Compared with the blank group, protein expression ratios p-PKA/PKA and p-CREB/CREB, as well as Bcl-2 levels, were significantly decreased in the model group (P<0.05). Compared with the model group, these indices were significantly increased in the medium-dose formula group (P<0.01). See Figure 5 and Table 7. (Image converted to text summary.)
3.9 Effects of Bushen Kaixuan Tongluo Formula on mRNA expression of PKA, CREB, and Bcl-2 in renal tissue of db/db mice
Compared with the blank group, mRNA expression of PKA, CREB, and Bcl-2 was significantly decreased in the model group (P<0.01). Compared with the model group, mRNA expression of PKA, CREB, and Bcl-2 was significantly increased in the medium-dose formula group (P<0.05). See Table 8.
Mechanistic context and discussion excerpt
cAMP signaling is a central hub for energy metabolism and gene regulation, crucial for renal homeostasis. Adenylate cyclase (AC) generates cAMP, which allosterically activates PKA. Activated PKA phosphorylates CREB to form p-CREB, which translocates into the nucleus, binds cAMP response elements (CRE) in target promoters, and initiates transcription of downstream effectors such as Bcl-2 to exert anti-apoptotic effects and mitigate cellular injury.
In DKD, chronic hyperglycemia suppresses AC activity and enhances phosphodiesterase (PDE) degradation, reducing cAMP levels, weakening PKA/CREB signaling, impairing mitochondrial function, antioxidant synthesis, and anti-apoptotic factor expression, thereby accelerating glomerulosclerosis and tubulointerstitial fibrosis.
Activating cAMP/PKA signaling helps maintain podocyte slit diaphragm proteins, protects the filtration barrier, reduces proteinuria, induces antioxidant genes (e.g., SOD2), and elevates Bcl-2 to curb ROS accumulation and protect cells under high-glucose conditions.
Normalizing blood glucose and improving lipid profiles can reduce AGEs accumulation, lessen GBM thickening and podocyte injury, and suppress renal lipid deposition and macrophage infiltration, ultimately delaying sclerosis and fibrosis.
In this study, the formula improved overall condition, significantly ameliorated renal function, and corrected glucose-lipid metabolism disorders in db/db mice. Histopathology showed relief of GBM thickening, mesangial expansion, and interstitial collagen deposition. Mechanistically, the formula upregulated renal PKA, CREB, and Bcl-2 mRNA and increased p-PKA/PKA, p-CREB/CREB, and Bcl-2 protein levels, activating core nodes of the cAMP pathway, enhancing antioxidative capacity, and improving filtration barrier function.
Conclusion: Bushen Kaixuan Tongluo Formula improves hepatic and renal function, corrects lipid and glucose metabolism disturbances, and attenuates renal fibrosis and oxidative stress via upregulation of the cAMP pathway, thereby conferring renoprotection. While multi-level evidence supports cAMP pathway involvement, downstream effectors such as BDNF were not examined and warrant future investigation.
Conflict of interest
No conflicts of interest.
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