**4.1 Statistical analysis**

Data obtained from this study are expressed as the means ± SEM. Statistical analyses were performed using GraphPad Prism 3.0 (GraphPad Software, Inc., San Diego, CA). Differences in blood pressure, serum creatinine, blood urea nitrogen, 24h urine protein and Urine TGF– β at different time points (weeks 0 to 8) within the groups, and differences of Ang II and

Molecular Mechanisms of Nephro-Protective

treatment

Before

Action of HE-86 Liquid Extract in Experimental Chronic Renal Failure 181

sham 137.3114.72 139.1314.06 125.507.15 150.5613.97 129.6329.16 control 140.5023.55\* 212.4643.26 199.9223.55 156.3320.72 202.4415.09 treatment 141.7726.45\* 148.5038.82## 152.4629.54## 141.0014.73# 176.0030.70# Table 4. Systolic blood pressure. Data represent the means ± SEM for groups of twelve rats treated with either HE-86 or empty vector (#P<0.05,##P<0.01,when compared against empty

vector-treated controls;\*P<0.05,\*\*P<0.01, when compared to normal sham-controls).

uraemic rats, manifesting HE-86 to attenuate the development of glomerular sclerosis.

Table 5. Effect of HE-86 liquid extract on urine TGF-β excretion in 5/6 nephrectomy in rats. (#P<0.05, ##P<0.01,when compared against empty vector-treated controls; \*P<0.05,

**9. Effects of HE-86 administration on localization of NF–κB in renal tissue** 

Immunohistochemical analysis was performed to determine the localization of NF–κB in the

response, has been shown to play a central role in inflammatory diseases, including kidney diseases [33]. In normal rats, only tubular epithelial cells were weakly stained by the monoclonal anti-NF–κB antibody, while glomeruli were hardly stained. In control uraemic rats, however, proximal tubular epithelial cells, especially of dilated tubules, were intensively stained by the anti-NF–κB antibody. In contrast, in the HE-86-treated uraemic

that in the control uraemic rats. The staining of NF–κB as shown in the control uraemic rats found increased NF–κB -positive (intensively stained) area in the renal cortex, whereas HE-86-treated rats showed markedly decreased NF–κB -positive area as compared to the control uraemic rats. These data demonstrate that HE-86 markedly reduces the overexpress of NF–

B, a critical transcriptional factor for controlling inflammatory

B in tubular epithelial cells was less prominent as compared with

High excreation of urine TGF-β1, which express both glomerular and tubulointerstitial injuries. To demonstrate further the anti-inflammatory effect of HE-86 on rat chronic renal failure, we determined the TGF-β1 levels within the urine by ELISA. Results demonstrated that compared with vehicle, He-86 treatment significantly reduced urinary TGF-β1 levels, corrected by decrease level of serum creatinine, throughout the entire disease course (P<0.05), indicating that HE-86 treatment may primarily suppress the local immune and inflammatory response within the diseased kidney. In contrast, overexpression of urine TGF-β1 was found in control uraemic rats as compared with normal rats (Table 5). The experimental result showed the administration of HE-86 significantly inversed high expression of urine TGF-β in

**8. Effects of HE-86 administration on urine TGF**-**β1** 

N Urine TGF-β(ug/L)

sham 12 1.830.64 control 12 1.900.56\* treatment 12 1.770.43#

\*\*P<0.01, when compared to normal sham-controls)

renal cortex (Fig.1-2). NF-

rats activation of the NF-

κB on the remnant tubular cells.

After treatment Second week Forth week Sixth week Eighth week

AT1R activation, TNF–α expression and NF-κb accumulation in sham, control and HE86 treated animals were assessed by one-way analysis of variance, followed by t-test. Results were considered statistically significant when the P value was <0.05.
