ß-amyloid peptide binds and regulates ectopic ATP synthase α-chain on neural surface.

Accumulation of the amyloid ß protein (Aß) in the brain is an important step in the pathogenesis of Alzheimer's disease. Many molecules could bind with Aß, among which some molecules mediate Aß neuronal toxicity. Thus, it is of interest to study the binding proteins of Aß, and the functions that might be affected by Aß. In the present study, we observed that accumulation of α-subunit of ATP synthase is associated with aggregates of Aß proteins in amyloid plaques of amyloid precursor protein/presennillin-1 transgenic mice, and identified the α-subunit of ATP synthase as one of the Aß binding proteins on the plasma membrane of neural cells by Western blot and mass spectrometry. In order to evaluate the consequences of the interaction between Aß and surface α-subunit of ATP synthase, the extracellular ATP generation was analyzed, which showed that aggregated Aß partially inhibited the extracellular generation of ATP, but was unable to significantly induce a decrease in cell surface ATP synthase α on neurons. These results suggest that the cell surface ATP synthase α is a binding protein for Aß on neural cells, the functional inhibition of surface ATP synthase might be involved in machinery of brain malfunction in Aß-mediated pathogenesis of Alzheimer's disease.

[To investigate the strategy of Chinese medicine for prevention and treatment of atherosclerosis based on vascular aging].

Atherosclerosis, a chronic degenerative disease mainly attacks the middle-aged and the aged population as they grow old. Anti-angiocellular aging has gradually become a new strategy for atherosclerosis. In the process of atherosclerosis developing, endothelial cell renewing is speeding. Various biological function disorders that induce blood vessel aging emerge, which leads to changes of the telomere and telomerase, resulting in aged endothelial cells and dysfunction. Telomere and telomerase may play key roles in the etiological factors such as inflammation and AS plaque. In our previous work we have found that Chinese compounds with Shen invigorating effects could not only obviously ameliorate the symptoms and functions of the senility, but also show significant effects on restraining atherosclerosis. We should actively study the mechanisms of Chinese medicine for treating atherosclerosis from Shen, and the mechanisms of Shen invigorating compounds for regulating angiocellular aging through the telomere pathway, thus providing evidence for establishing vascular cell aging based atherosclerosis prevention and treatment strategies by Chinese medicine.

[Dynamic proteomic analysis of liver tissue in the carbon tetrachloride-induced liver fibrosis rat model].

OBJECTIVE: To elucidate the dynamic physiopathologic mechanisms of liver fibrosis by investigating the differential proteome of liver tissue during progression of liver fibrosis in a chemically induced rat model. METHODS: Following treatment with carbon tetrachloride (CCl4), livers were harvested from rats at various time points. The respective total protein extracts were resolved by two-dimensional gel electrophoresis (2-DE) and compared to identify differentially expressed protein spots, which were then analyzed by matrix-assisted laser desorption/ionization two-stage time-of-flight mass spectrometry (MALDI-TOF/TOF-MS) and identified by database querying. The differential expression of selected proteins was validated by Western blotting and immunohistochemical methods. Statistical analyses were carried out by the least significant difference method of one-way ANOVA for parametric data or by the H test for non-parametric data. RESULTS: The severity scores of liver fibrosis increased in a time-dependent manner following CCl4 exposure (post-induction weeks: 3 less than 6 less than 9). Forty-four protein spots were different on the 2-DE maps for the different time points, among which the CK8 and CK18 proteins were identified and verified as significantly differentially expressed as liver fibrosis progressed. Protein expressions of CK8/CK18 were enhanced upon CCl4 exposure and increased over time (untreated controls: 0.113 ± 0.005/0.170 ± 0.030; CCl4-induced rats at week 3: 0.473 ± 0.046/0.530 ± 0.070, at week 6: 0.682 ± 0.087/0.780 ± 0.080, and at week 9: 0.837 ± 0.096/1.390 ± 0.130). Moreover, the rate of "a" determinant mutations for CK8/CK18 was also significantly differently between weeks 3, 6, and 9 (F = 196.085/74.088, 13.870/16.115, and 75.800/75.900, P less than 0.01). CONCLUSION: Dynamic proteomic analysis of liver tissue can indicate physiopathologic changes in protein expressions that are related to liver fibrosis induced by CCl4. Proteins with differential expression in CCl4-damaged fibrotic liver are associated with cell growth, development and differentiation, cell proliferation and apoptosis, angiogenesis or reconstitution, oxidative stress, substance metabolism and transport, and signal transduction.

Symptoms Based on Deficiency Syndrome in Traditional Chinese Medicine Might Be Predictor of Frailty in Elderly Community Dwellers.

BACKGROUND: The most widely used frailty phenotype and frailty indexes are either time-consuming or complicated, thus restricting their generalization in clinical practice; and therefore, an easier and faster screening tool is needed to be developed. OBJECTIVE: To select sensitive symptoms in traditional Chinese medicine (TCM) and study whether they can improve the risk prediction of frailty. METHODS: This is a cross-sectional study enrolling 2249 Chinese elderly community dwellers. Data were collected via face-to-face inquiries, anthropometric measurements, laboratory tests, and community health files. Frailty was the main outcome measure, and it was evaluated by Fried's frailty phenotype (FP). The ordinal logistic regression model was used to identify the factors associated with frailty. The risk assessment plot was used to compare the discriminative ability for frailty among models with and without TCM symptoms. RESULTS: The identified sensitive influential factors for frailty included age, education level, dietary habits, chronic obstructive pulmonary disease, diabetes, cerebral infarction, osteoporosis, cold limbs, lethargy and laziness in speaking and moving, weakness of lower limbs, slow movement, dry mouth and throat, and glazed expression. The risk prediction for "frailty cumulative components ≥1" was not significantly increased, while for "frailty cumulative components ≥2", a new model developed with the above selected TCM symptoms had a higher AUC than the baseline model without it (0.79 VS 0.81, P=0.002). And the NRI and IDI for the new model were 41.4% (P=0.016) and 0.024% (P=0.041), respectively. CONCLUSION: This research might provide an easier and faster way for early identification and risk prediction of frailty in elderly community dwellers.

[Mechanism of hepatocytes transdifferentiation to bile duct epithelial cells and intervention of huangqi decoction].

OBJECTIVE: To investigate the mechanism of hepatocytes transdifferentiation to bile duct epithelial cells (BECs) and intervention of Huangqi decoction (HQD) on hepatic fibrosis formation in rats with secondary cholestasis. METHODS: Seventy-five SD male rats were made into cholestatic hepatic fibrosis model animals by bile duct ligation, and randomized into the control group (n = 50) and the HQD group (n = 15). Starting from one week after modeling, they were administered orally with saline and HQD respectively for four weeks. Besides, a sham-operated group was set up with 10 rats operated by choledochus segregating only and administered after then with saline. Rats were killed in batches at different time points, i.e. each five from the control group and sham-operated group at the end of the 1st week, five from the control group for each time at the end of the 2nd, 3rd and 4th week, and all the remaining rats at the end of the 5th week. Their liver tissues were taken for histological change examination, content of hydroxyproline (Hyp) determination; protein expression of BECs marker cytokeratin 7 (CK7) and the hepatocyte specific antigen HepPar detection by Western blot, and CK7-Hep Par co-localization by laser confocal microscopy. Then IPP software was used to analyze Sirius red stained positive areas of CK7 and Hep Par, as well as the average IOD of CK7/Hep Par co-localization. RESULTS: Hepatocytes in hepatic tissues (Hep Par positive cell) in the model rats decreased gradually along was time went by after modeling (Sham > M1w > M2w > M3w > M4w > M5w), which was in parallel with the increase of BECs (CK7 positive cells), degree of fibrosis, Hyp content and CK7 protein expression. Increasing of co-localized positive cells of CK7/Hep Par began at 1 week and reached the peak 3 weeks after modeling, then it decreased gradually. The Hep Par protein expression was negatively correlated with that of CK7; the Hep Par positive cell expression was negatively correlated with CK7 positive cell expression and collagen deposition; while the CK7 positive cell expression was positively correlated with the collagen deposition in the liver tissue. Compared with the model control group, the mortality, CK7/Hep Par co-localized positive cells, fibrosis degree, Hyp content and CK7 protein expression were lesser obviously (P < 0.01), while Hep Par positive cell and protein expressions were higher significantly in the HQD group. CONCLUSIONS: Hepatocytes transdifferentiation to BECs might be a key pathological element for secondary cholestatic hepatic fibrosis formation; the restraining action of HQD is possibly a major action mechanism of HQD for effectively intervening and treating secondary cholestasis hepatic fibrosis.

[Effects of Yiguanjian Decoction on liver cirrhosis formation:a differential proteomics study in rats].

OBJECTIVE: To investigate the effects of Yiguanjian Decoction, a compound traditional Chinese herbal medicine, on rats with cirrhosis based on the method of differential proteomics. METHODS: Wistar male rats (n=48) were randomly divided into normal control group (n=12) and model-making group (n=36). Rat cirrhosis model was established by intraperitoneal injection of 50% carbon tetrachloride (CCl4) plus olive oil solution (1 mL/kg, twice weekly for 9 weeks). After 3- and 6-week injection, 6 rats each time were sacrificed for dynamic observation before medicine intervention, and the 24 remained rats were randomly divided into untreated group (n=12) and Yiguanjian Decoction group (n=12) at the first day of the 7th week. All animals were sacrificed by the end of the 9th week, and total protein of liver tissue was isolated by two-dimensional gel electrophoresis (2-DE); some differentially expressed protein spots were analyzed and identified by matrix-assisted laser desorption/ionization two-stage time-of-flight mass spectrometry (MALDI-TOF/TOF-MS) and database querying. Protein expressions of Cu/Zn superoxide dismutase (Cu/Zn SOD) and DJ-1 were validated by Western blot and immunohistochemical methods. RESULTS: 2-DE maps with high resolution and good repeatability were obtained. In all 50 protein spots identified by MALDI-TOF/TOF-MS and database querying, there were 5 protein spots related to oxidative stress named Cu/Zn SOD, DJ-1, glutathione synthetase, glutathione S-transferase Yb-1 subunit and aldo-keto reductase family 7, A2 respectively. Compared with the normal control group, expressions of Cu/Zn SOD, DJ-1, glutathione S-transferase Yb-1 subunit and aldo-keto reductase family 7, A2 in the untreated group were decreased significantly. Expressions of Cu/Zn SOD and aldo-keto reductase family 7, A2 were decreased time-dependently. Compared with the untreated group in 9th week, protein expressions of Cu/Zn SOD, DJ-1, glutathione S-transferase Yb-1 subunit and aldo-keto reductase family 7, A2 in the Yiguanjian Decoction groups were increased significantly while expression of glutathione synthetase was decreased notably. Western blot and immunohistochemical results of Cu/Zn SOD and DJ-1 expressions coincided with proteomics results. CONCLUSION: Anti-oxidative depression is a key pathological change of cirrhosis induced by CCl4 in rats, and increasing expression of proteins related to anti-oxidative stress may be a major mechanism of Yiguanjian Decoction in treating cirrhosis induced by CCl4 effectively.

Effect of Quercetin on Atherosclerosis Based on Expressions of ABCA1, LXR-α and PCSK9 in ApoE-/- Mice.

OBJECTIVE: To investigate the effect of quercetin on ATP binding cassette transporter A1 (ABCA1), liver X receptor (LXR), and proprotein convertase subtilisin/kexin type 9 (PCSK9) expressions in apoE-knockout (ApoE-/-) mice. METHODS: The high-fat diet-induced atherosclerosis (AS) in ApoE-/- mice was established. Thirty-six mice were divided into 3 groups using random number table method: model group (n=12), quercetin group (n=12), and atorvastatin group (n=12), with C57BL/6J mice of the same strain and age as the control group (n=12). Quercetin group and atorvastatin group were administrated with quercetin and atorvastatin by oral gavage, with doses of 12.5 and 4 mg/(kg•d), respectively. Animals in the control and model groups were given an equal volume of distilled water by oral gavage once per day for a total of 12 weeks. Western blot and immunohistochemical methods were employed to determine the aortic ABCA1, LXR-α and PCSK9 protein expression. Enzyme linked immunosorbent assay method was used to detect the expression of serum total cholesterol (TC), triglyceride (TG), high density lipoprotein-cholesterol (HDL-C), low density lipoprotein-cholesterol (LDL-C), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IL-10, combined with tissue pathological examination. RESULTS: ApoE-/- mice fed with a high-fat diet had notable atherosclerosis lesions, with reduced ABCA1, LXR-α and IL-10 levels (all P<0.01), elevated PCSK9, TNF-α and IL-6 expression, and increased TC and LDL-C contents (all P<0.01). After quercetin intervention, the areas of AS plaques and the expressions of PCSK9, TNF-α and IL-6 were significantly reduced (all P<0.01), while the expressions of ABCA1 and LXR-α were increased significantly (all P<0.01). CONCLUSION: Quercetin effectively interfered with AS development by regulating the expressions of ABCA1, LXR- α and PCSK9 in ApoE-/- mice.

Tiaoxin Recipe, a Chinese herbal formula, inhibits microRNA-34a expression in the APPswe/PS1ΔE9 mouse model of Alzheimer's disease.

OBJECTIVE: To investigate the effect and underlying mechanisms of Tiaoxin Recipe (a Chinese herbal formula) treatment on Alzheimer's disease (AD). METHODS: Twelve-week-old APPswe/PS1ΔE9 (APP/PS1) double transgenic mice were used as a model of AD-afflicted mice. One group of mice was treated with Tiaoxin Recipe by gastrogavage for 12 weeks, while two other groups were given intraperitoneal injections of nicotinamide adenine dinucleotide or FK866 for 4 weeks. Morris water maze and thioflavin S staining tests were performed to evaluate cognitive impairment and amyloid plaque deposition, respectively. Serum amyloid-ß1-42 (Aß1-42) content was detected using an enzyme-linked immunosorbent assay, and quantitative reverse transcription-polymerase chain reaction was performed to examine the expression levels of microRNA-34a (miR-34a) in cortex and hippocampus samples of the study mice. RESULTS: Compared with the normal control group, the memory and learning abilities of the APP/PS1 model group were found to be impaired (P < 0.01), as shown by the increased levels of senile plaque deposition in cortex and hippocampus (P < 0.01), miR-34a expression (P < 0.01) and serum Aß1-42 content (P < 0.01). Treatment with Tiaoxin Recipe significantly reduced memory impairment (P < 0.01) by reducing amyloid plaque accumulation in cortex and hippocampus (P < 0.01), miR-34a expression (P < 0.01) and serum Aß1-42 content (P < 0.01) in APP/PS1 mice. CONCLUSION: Tiaoxin Recipe is a viable complementary or alternative therapeutic treatment that is capable of delaying the development of early-stage AD by inhibiting the expression of miR-34a.

A target role for mast cell in the prevention and therapy of hepatic fibrosis.

Hepatic fibrosis is a common pathological process of chronic hepatic disease. Despite the extensive studies, the pathophysiological mechanisms in hepatic fibrosis remain unclear. Mast cell has a variety of physiological and pathological functions through the production of heparin, histamine, neutrophil chemoattractants, immunoregulatory cytokines, and mast cell-specific serine proteases tryptase and chymase. The survival and proliferation of mast cell are dependent upon stem cell factor. More recently, the data have suggested that mast cell has been associated with hepatic fibrosis in many chronic liver diseases. However, to what extent the mast cell effects the hepatic fibrosis remains to be clarified. Several therapeutic approaches to inhibit mast cell activation have already demonstrated some clinical utility in tissue fibrosis or inflammatory diseases such as the use of mast cell stabilizers, inhibitors of tryptase or chymase, blockade of stem cell factor and anti-IgE therapy. The article introduces the hypothesis that mast cell has a central role when it is affected by its activation state in the progression of hepatic fibrosis, thus new therapeutic strategies for treatment of hepatic fibrosis are suggested by this hypothesis. Considering the important role of mast cell and the development of these tangible therapeutic approaches in hepatic fibrosis will enable us to target any types of chronic liver diseases, which appears to be a more reasonable or a promising strategy.

Effect of qi-tonifying and stasis-eliminating therapy on expression of vascular endothelial growth factor and its receptors Flt-1, Flk-1 in the brain of intracerebral hemorrhagic rats.

OBJECTIVE: To investigate the effects and mechanism of qi-tonifying and stasis-eliminating (QTSE) therapy on the expression of vascular endothelial growth factor (VEGF) and its receptors Flt-1 and Flk-1 in the brains of intracerebral hemorrhagic (model) rats. METHODS: One hundred and eighty Sprague-Dawley rats were randomly divided into six groups: the normal group (n=5), the sham-operative (SO) group (n=35), the model group (n=35), the QTSE group (n=35), the QT group (n=35) and the SE group (n=35). All the rats except those in the normal group and SO group were established into an intracerebral hemorrhage(ICH) model by intracerebral injection of collagenase type VII and the latter three were orally administered with Buyang Huanwu Decoction (a classical recipe for QTSE) or with some of its components for qi-tonification and for stasis-elimination, respectively. To the other three groups, normal saline solutions were given instead. Behavioral tests were carried out in the animals randomly chosen from each group on days 1, 2, 4, 7, 14, 21 and 28 after modeling. The expressions of VEGF, Flk-1 and Flt-1 were determined by immunohistochemistry and the number of vascular segments with positive expression in the injured brain area of the rats was calculated. RESULTS: From day 7 onwards, the asymmetric forelimb use rate in the QTSE group recovered more significantly than that in the other model groups. In the model group, the expressions of VEGF, Flk-1 and Flt-1 appeared on day 1 and reached a peak on day 21, then weakened gradually. In the QTSE group, as compared with the other model groups, a higher level of VEGF expression was shown from day 7 (P<0.01) and a higher level of Flt-1 expression was shown from the 7th day to the 21st day (P<0.01). CONCLUSION: QTSE therapy can up-regulate the expressions of VEGF and its receptors (Flk-1 and Flt-1) and improve the recovery of kinetic function in the ICH rats, which may be correlated with its action in modulating vascular regeneration to promote the reconstruction of microvascular networks in the damaged areas.

[Exploration on correlation of syndrome proteomics and syndromatology of TCM].

The advent of the Post-Human Genome Project Era, represented by the raising of proteomics, would inevitably lead to the change of molecular biology from topical view to holistic with its thought turning from linear to complex mode. Based on the proteomics development in recent years, the authors summarized the methodology of TCM syndromatologic research, advocated in using two-dimensional gel electrophoresis (2-DE) and bioinformatics to identify different proteins. Proteomics should be led into the research of TCM syndrome categorization and the rule of evolution, which is necessary for researching the integration of the TCM study with proteomics and even with modern molecular biology based on molecular epidemiology level. Owing to the gradually developed coherence and mutual penetration of proteomics and TCM on the thinking method in studying life science, it has denoted the necessity and importance of integration of TCM and Western medicine in investigating the complex vital life phenomena.

[Nuclear factor-kappaB mRNA and protein expression in stomach tissue of rats with gastric ulcer recurrence and effect of jianwei yuyang granule on its expression].

OBJECTIVE: To observe the inflammatory reaction, nuclear factor-kappaB (NF-kappaB) mRNA and protein expression in stomach tissue of rats with gastric ulcer recurrence and the effect of Jianwei Yuyang granule (JYG) on them. METHODS: Gastric ulcer and its recurrent lesion were successively induced by acetic acid and interliukin1-beta (IL-1beta), and the model rats were divided into the sham operation group, the model group, the omeprazole (correction of omepraxole) group and the JYG group to observe the state of chronic inflammatory cell, neutrophil count, NF-kappaBmRNA and protein expression in stomach tissue. RESULTS: On the 16th and 92th day after administration, the increase of chronic inflammatory cell, neutrophil, NF-kappaBmRNA and protein expression in the model group was more significant than those in the sham operated group (P < 0.01), while that was lower in the JYG group than in the model group (P < 0.05, P <0.01), but with no remarkable difference to the omepraxole group. In addition, the mRNA and protein expression of NF-kappaB were correlated closely with the count of chronic inflammatory cell and neutrophil respectively (P < 0.01). CONCLUSION: NF-kappaB may play an important role in regulating inflammatory reaction during the healing and recurrence processes of gastric ulcer induced by acetic acid. JYG may suppress inflammatory reaction by inhibiting the activation and expression of NF-kappaB in stomach tissue, which may be one of the mechanisms of JYG in preventing the recurrence of gastric ulcer.

[Effects of jianwei yuyang granule on inflammatory reaction and NF-kappaB expression in rat gastric mucosa of ulcer healing and recurrence].

OBJECTIVE: To observe the effects of Jianwei Yuyang granule (JWYY) on inflammatory reaction and NF-kappaB expression in rat gastric mucosa of ulcer healing and recurrence. METHOD: Gastric ulcer was induced in rat by acetic acid according Okeba's method with some modification and the recurrence model was induced by IL-1beta. Pathohistology of ulcer healing and recurrence was observed. Density of inflammatory cell infiltrating regenerative mucosa, NF-kappaB protein and mRNA expression were measured. RESULT: JWYY had effects on improving the quality of ulcer healing, reducing the rate of ulcer recurrence, decreasing the density of inflammatory cell infiltrating regenerative mucosa and suppressing the activation and expression quantity of NF-kappaB protein and mRNA. CONCLUSION: JWYY may promote the ulcer healing and prevent the recurrence of the gastric ulcer by suppressing the activation of NF-kappaB and the following inflammatory reaction.

Effect of atorvastatin on expression of TLR4 and NF-κB p65 in atherosclerotic rabbits.

OBJECTIVE: To study the effect of atorvastatin on atherosclerotic rabbits. METHODS: A total of 60 New Zealand male rabbits were randomly divided into the normal group, model group and atorvastatin group. The replication rabbit atherosclerotic model with immune injury combined with a high fat diet feeding was used. All rabbits were sacrificed after 3 months. TLR4 and NF-κB p65 were observed by HE staining, immunohistochemistry and western blotting. RESULTS: The expression of TLR4, NF-κB p65 were significantly increased in the model group compared with the normal group. The expression of TLR4 and NF-κB p65 decreased significantly in the atorvastatin group, and there was no difference compared with the normal group. CONCLUSIONS: The effect of atorvastatin on atherosclerosis may be achieved by the inhibition of the expression of TLR4 and NF-κB p65.