Table 1.
Nucleotide sequence of primers used in Real-time PCR.
Table 2.
Effect of a combination of Rhizoma Coptidis alkaloids, Radix et Rhizoma Rhei polysaccharides, and Radix Scutellaria flavones (APF) on glucose and lipid metabolism, urinary albumin excretion and kidney function and renal function in db/db mice.
Fig 1.
Effect of a combination of Rhizoma Coptidis alkaloids, Radix et Rhizoma Rhei polysaccharides, and Radix Scutellaria flavones (APF) on renal histopathology and ultrastructural pathology.
(a-e) hematoxylin and eosin (HE) stain. (f-j) Periodic Acid Schiff (PAS) stain. Original magnification (a–j) × 400. (k-o) Electron microscopy (EM) analysis, Representative images of glomerular basement membrane thickening and mesangial matrix expansion, scale bars 2 μm, original magnification electron microscopy × 6000. (p) Ratio of the mesangial matrix area to total glomerular area (M/G) in PAS staining. Data are expressed as mean ± S.D., n = 10, **p < 0.01 as compared with db/db group.
Fig 2.
Effect of a combination of Rhizoma Coptidis alkaloids, Radix et Rhizoma Rhei polysaccharides, and Radix Scutellaria flavones (APF) on renal inflammation in db/db mice.
(a-b) Western blot analysis of protein levels; (c-d) Real-time RCR analysis of mRNA levels; ICAM-1: intercellular adhesion molecule-1; MCP-1: monocyte chemotactic protein-1; TNF-α: tumor necrosis factor-α; IL-1β: interleukin-1β. Data are expressed as mean ±S.D., n = 3 for Western blot, and n = 5 for Real-time PCR, *p<0.05, **p<0.01 as compared with db/db group.
Fig 3.
Effect of a combination of Rhizoma Coptidis alkaloids, Radix et Rhizoma Rhei polysaccharides, and Radix Scutellaria flavones (APF) on renal nuclear factor-κB (NF-κB) signaling pathway.
IKKα: inhibitor of nuclear factor-κB kinase subunit α; IκBα, inhibitor of nuclear factor-κB subunit α; p-IκBα: phospho-IκBα; NF-κBp65: nuclear factor-κBp65; Data are expressed as mean ±S.D., n = 3, *p<0.05, **p<0.01 as compared with db/db group
Fig 4.
Effect of a combination of Rhizoma Coptidis alkaloids, Radix et Rhizoma Rhei polysaccharides, and Radix Scutellaria flavones (APF) on renal fibrosis.
(a-e) Masson’s modified trichrome histological (Masson); (p) Ratio of area with collagen accumulation to total glomerular area; (f-j) Immunohistochemistry of collagen I; (k-o) Immunohistochemistry of collagen IV. Original magnification (a–o) × 400; (q) Quantitative analysis of immunohistochemical staining of collagen I (Col I); (r) glomerular of collagen IV (Col IV); (s) interstitial of collagen IV (Col IV); (t-u) Real-time RCR analysis of collagen I and collagen IV mRNA levels. Data are expressed as mean ±S.D., n = 5, **p<0.01 as compared with db/db group.
Fig 5.
Effect of a combination of Rhizoma Coptidis alkaloids, Radix et Rhizoma Rhei polysaccharides, and Radix Scutellaria flavones (APF) on renal TGF-β1 and its receptor expression.
(a-e) Immunohistochemistry of TGF-β1; (f) Quantitative analysis of immunohistochemical staining of TGF-β1; (g-h) Western blot analysis of TGF-β1 and α-SMA protein levels; (i-j) Real-time RCR analysis of TGF-β1 and TβRⅡmRNA levels. a: db/m, b: db/db, c: APF 300mg/kg, d: 600 mg/kg, e: metformin. Data are expressed as mean ±S.D., n = 3 for Western blot, and n = 5 for Immunohistochemistry and Real-time PCR, *p<0.05, **p<0.01 as compared with db/db group.
Fig 6.
Effect of a combination of Rhizoma Coptidis alkaloids, Radix et Rhizoma Rhei polysaccharides, and Radix Scutellaria flavones (APF) on renal TGF-β1/Smad signaling pathway.
(a-c) Western blot analysis of Smad7, p-Smad3 and p-Smad2 protein levels; d) Real-time RCR analysis of Smad7 mRNA level. Data are expressed as mean ±S.D., n = 3 for Western blot, and n = 5 for Real-time PCR, *p<0.05, **p<0.01 as compared with db/db group.
Fig 7.
Effects of a combination of Rhizoma Coptidis alkaloids, Radix et Rhizoma Rhei polysaccharides, and Radix Scutellaria flavones (APF) on cell proliferation and fibrosis in mesangial cells incubated at high glucose.
NG: normal glucose; HG: high glucose; APF 4, 8, and 16 μg /mL represent mesangial cells incubated in high glucose concentration treated with 4, 8, and 16 μg /mL APF, respectively. (a-d) Western blot analysis of α-SMA, CollagenⅠ, TGF-β1 and NF-κB protein levels; (e) Cell proliferation. Data are expressed as mean ±S.D., n = 5 for cell proliferation, and n = 3 for Western blot, *p<0.05, **p<0.01, compared to control cells incubated at high glucose concentration with no APF.