AIM: To evaluate the effect of Chinese traditional medicinal prescription, JIANPI HUOXUE decoction (JHD) on cytokine secretion pathway in rat liver induced by lipopolysaccharide (LPS).

METHODS: Twenty-four male SD rats were divided into normal group (n = 4), model group (n = 10) and JHD group (n = 10) randomly. Rats in model group and JHD group were administrated with normal saline or JHD via gastrogavage respectively twice a day for 3 d. One hour after the last administration, rats were injected with LPS via tail vein, 50 μg/kg. Simultaneously, rats in normal group were injected with equivalent normal saline. After LPS stimulation for 1.5 h, serum and liver tissue were collected. Pathological change of liver tissues was observed through hematoxylin-eosin (H.E.) staining. Tumor necrosis factor alpha (TNF-α) in serum were assayed by enzyme linked immunosorbent assay (ELISA). The protein expression of TNF-α, phosphorylated inhibit-κB (p-IκB) and CD68 in liver were assayed by Western blot. The distribution of CD68 protein in liver was observed through immunohistochemical staining. The mRNA expression of TNF-α, interleukin-6 (IL-6), CD14, toll-like receptor 2 (TLR2) and TLR4 in liver were assayed by real-time RT-PCR.

RESULTS: Predominant microvesicular change, hepatocyte tumefaction and cytoplasm dilution were observed in liver tissues after LPS administration as well as obvious CD68 positive staining in hepatic sinusoidal. After LPS stimulation, serum TNF-α (31.35 ± 6.06 vs 12 225.40 ± 9007.03, P < 0.05), protein expression of CD68 (1.13 ± 0.49 vs 3.36 ± 1.69, P < 0.05), p-IκB (0.01 ± 0.01 vs 2.07 ± 0.83, P < 0.01) and TNF-α (0.27 ± 0.13 vs 1.29 ± 0.37, P < 0.01) in liver and mRNA expression of TNF-α (1.96 ± 2.23 vs 21.45 ± 6.00, P < 0.01), IL-6 (4.80 ± 6.42 vs 193.50 ± 36.36, P < 0.01) and TLR2 (1.44 ± 0.62 vs 4.16 ± 0.08, P < 0.01) in liver were also increased significantly. These pathological changes were all improved in JHD group. On the other hand, TLR4 mRNA (1.22 ± 0.30 vs 0.50 ± 0.15, P < 0.05) was down-regulated and CD14 mRNA increased but not significantly after LPS stimulation.

CONCLUSION: JHD can inhibit cytokine secretion pathway induced by LPS in rat liver, which is probably associated with its regulation on CD68, p-IκB and endotoxin receptor TLR2.

Kudzu (Pueraria lobata) is one of the earliest medicinal plants used to treat alcohol abuse in traditional Chinese medicine for more than a millennium. However, little is known about its effects on chronic alcoholic liver injury. Therefore, the present study observed the effects of puerariae radix extract (RPE) on chronic alcoholic liver injury as well as Kupffer cells (KCs) activation to release tumor necrosis factor alpha (TNF-α) induced by gut-derived endotoxin in rats and macrophage cell line. RPE was observed to alleviate the pathological changes and lipids deposition in liver tissues as well as the serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and hepatic gamma-glutamyl transpeptidase (GGT) activity. Meanwhile, RPE inhibited KCs activation and subsequent hepatic TNF-α expression and downregulated the protein expression of endotoxin receptors, lipopolysaccharide binding protein (LBP), CD14, Toll-like receptor (TLR) 2, and TLR4 in chronic alcohol intake rats. Furthermore, an in vitro study showed that RPE inhibited the expression of TNF-α and endotoxin receptors, CD14 and TLR4, induced by LPS in RAW264.7 cells. In summary, this study demonstrated that RPE mitigated liver damage and lipid deposition induced by chronic alcohol intake in rats, as well as TNF-α release, protein expression of endotoxin receptors in vivo or in vitro.

AIM: To investigate whether circulating microRNAs (miRNAs) can serve as molecular markers to predict liver injury resulted from chronic hepatitis B (CHB).

METHODS: The profiles of serum miRNA expression were first generated with serum samples collected from 10 patients with CHB and 10 healthy donors (Ctrls) by microarray analysis. The levels of several miRNAs were further quantitated by real-time reverse transcription polymerase chain reaction with serum samples from another 24 CHB patients and 24 Ctrls. Serum samples of 20 patients with nonalcohlic steatohepatitis (NASH) were also included for comparison. The comparison in the levels of miRNAs between groups (CHB, NASH and Ctrl) was analyzed with Mann-Whitney U-test. The correlation between miRNAs and clinical pathoparameters was analyzed using Spearman correlation analysis or canonical correlation analysis. The receiver-operator characteristic (ROC) curves were also generated to determine the specificity and sensitivity of each individual miRNA in distinguishing patients with CHB from Ctrls.

RESULTS: miRNA profile analysis showed that 34 miRNAs were differentially expressed between CHB and Ctrl subjects, in which 12 were up-regulated and 22 down-regulated in CHB subject (fold change > 2.0 and P < 0.01). The median levels of miR-122, -572, -575 and -638 were significantly higher (P < 1.00 × 10-5) while miR-744 significantly lower (P < 1.00 × 10-6) in CHB compared with the Ctrl. The levels of miR-122, -572 and -638 were also higher (P < 1.00 × 10-3) while the level of miR-744 lower in CHB (P < 0.05) than in NASH, although the difference between them was not as significant as that between CHB and Ctrl. ROC curve analysis revealed that the levels of miR-122, -572, -575, -638 and -744 in serum were sensitive and specific enough to distinguish CHB, NASH and Ctrl. Multivariate analysis further showed that the levels of these miRNAs were correlated with the liver function parameters. Most significantly, it was the scatter plot of principal component with the levels of these miRNAs, but not the parameters of liver function, which clearly distinguished CHB, NASH and Ctrl subjects.

CONCLUSION: Serum levels of miR-122, -572, -575, -638 and -744 are deregulated in patients with CHB or NASH. The levels of these miRNAs may serve as potential biomarkers for liver injury caused by CHB and NASH.

Traditional Chinese medicine (TCM) syndrome, also called ZHENG, is the basis concept of TCM theory. It plays an important role in TCM practice. There are excess and deficiency syndromes in TCM syndrome. They are the common syndromes in chronic hepatitis B (CHB) patients. Here we aim to explore serum protein profiles and potential biomarkers for classification of TCM syndromes in CHB patients. 24 healthy controls and two cohorts of CHB patients of excess syndrome (n = 25) or deficiency syndrome (n = 19) were involved in this study. Protein profiles were obtained by surface-enhanced laser desorption ionization time-flight mass spectrometry (SELDI-TOF/MS) and multiple analyses were performed. Based on SELDI ProteinChip data, healthy controls and CHB patients or excess and deficiency syndromes in CHB patients were obviously differentiated by orthogonal partial least square (OPLS) analysis. Two significant serum proteins (m/z 4187 and m/z 5032) for classifying excess and deficiency syndromes were found. Moreover, the area under the receiver operating characteristic (ROC) curve was 0.887 for classifying excess and nonexcess syndrome, and 0.700 for classifying deficiency and nondeficiency syndrome, respectively. Therefore, the present study provided the possibility of TCM syndrome classification in CHB patients using a universally acceptable scientific approach.

In Traditional Chinese Medicine (TCM), treatment based on ZHENG (also called TCM syndrome and pattern) differentiation has been applied for about 3 thousand years, while there are some difficulties to communicate with western medicine. In the present work, metabonomic methods were utilized to differentiate ZHENG types and evaluate the therapeutic efficiency of Fuzhenghuayu (FZHY) tablet in hepatitis-B-caused cirrhosis (HBC). Urine samples of 12 healthy volunteers (control group, CG) and 31 HBC patients (HBCG) were analyzed by gas chromatography mass spectrometry (GC/MS) and multivariate statistical analysis. The significantly changed metabolites between CG and HBCG were selected by PLS-DA loading plot analysis. Moreover, 4 ZHENGs were differentiated mutually, suggesting that there was urine metabolic material basis in ZHENG differentiation. The efficiency of FZHY tablet on subjects with spleen deficiency with dampness encumbrance syndrome (SDDES) and liver-kidney yin deficiency syndrome (LKYDS) was better than that of other syndromes. The efficiency of FZHY treatment based on ZHENG differentiation indicated that accurately ZHENG differentiating could guide the appropriate TCM treatment in HBC.

Background

Cancer stem cells (CSCs) play an important role in the development and recurrence of malignant tumors including glioma. Notch signaling, an evolutionarily conserved pathway mediating direct cell-cell interaction, has been shown to regulate neural stem cells (NSCs) and glioma stem cells (GSCs) in normal neurogenesis and pathological carcinogenesis, respectively. However, how Notch signaling regulates the proliferation and differentiation of GSCs has not been well elucidated.

Methods

We isolated and cultivate human GSCs from glioma patient specimens. Then on parallel comparison with NSCs, we inhibited Notch signaling using γ-secretase inhibitors (GSI) and assessed the potential functions of Notch signaling in human GSCs.

Results

Similar to the GSI-treated NSCs, the number of the primary and secondary tumor spheres from GSI-treated GSCs decreased significantly, suggesting that the proliferation and self-renewal ability of GSI-treated GSCs were attenuated. GSI-treated GSCs showed increased differentiation into mature neural cell types in differentiation medium, similar to GSI-treated NSCs. Next, we found that GSI-treated tumor spheres were composed of more intermediate progenitors instead of CSCs, compared with the controls. Interestingly, although inhibition of Notch signaling decreased the ratio of proliferating NSCs in long term culture, we found that the ratio of G2+M phase-GSCs were almost undisturbed on GSI treatment within 72 h.

Conclusions

These data indicate that like NSCs, Notch signaling maintains the patient-derived GSCs by promoting their self-renewal and inhibiting their differentiation, and support that Notch signal inhibitor GSI might be a prosperous candidate of the treatment targeting CSCs for gliomas, however, with GSI-resistance at the early stage of GSCs cell cycle.

Background

Huangqi decoction was first described in Prescriptions of the Bureau of Taiping People's Welfare Pharmacy in Song Dynasty (AD 1078), and it is an effective recipe that is usually used to treat consumptive disease, anorexia, and chronic liver diseases. Transforming growth factor beta 1 (TGFβ1) plays a key role in the progression of liver fibrosis, and Huangqi decoction and its ingredients (IHQD) markedly ameliorated hepatic fibrotic lesions induced by ligation of the common bile duct (BDL). However, the mechanism of IHQD on hepatic fibrotic lesions is not yet clear. The purpose of the present study is to elucidate the roles of TGFβ1 activation, Smad-signaling pathway, and extracellular signal-regulated kinase (ERK) in the pathogenesis of biliary fibrosis progression and the antifibrotic mechanism of IHQD.

Methods

A liver fibrosis model was induced by ligation of the common bile duct (BDL) in rats. Sham-operation was performed in control rats. The BDL rats were randomly divided into two groups: the BDL group and the IHQD group. IHQD was administrated intragastrically for 4 weeks. At the end of the fifth week after BDL, animals were sacrificed for sampling of blood serum and liver tissue. The effect of IHQD on the TGFβ1 signaling pathway was evaluated by western blotting and laser confocal microscopy.

Results

Decreased content of hepatic hydroxyproline and improved liver function and histopathology were observed in IHQD rats. Hepatocytes, cholangiocytes, and myofibroblasts in the cholestatic liver injury released TGFβ1, and activated TGFβ1 receptors can accelerate liver fibrosis. IHQD markedly inhibited the protein expression of TGFβ1, TGFβ1 receptors, Smad3, and p-ERK1/2 expression with no change of Smad7 expression.

Conclusion

IHQD exert significant therapeutic effects on BDL-induced fibrosis in rats through inhibition of the activation of TGFβ1-Smad3 and TGFβ1-ERK1/2 signaling pathways.