Cheng Xiao

Systems Biology Guided by Chinese Medicine Reveals New Markers for Sub-typing Rheumatoid Arthritis Patients

Background:Complex chronic diseases such as rheumatoid arthritis have become a major challenge in medicine and for the pharmaceutical industry. New impulses for drug development are needed. Objective:A systems biology approach is explored to find subtypes of rheumatoid arthritis patients enabling a development towards more personalized medicine. Methods:Blood samples of 33 rheumatoid arthritis (RA) patients and 16 healthy volunteers were collected. The RA patients were diagnosed according to Chinese medicine (CM) theory and divided into 2 groups, the RA Heat and RA Cold group. CD4+ T-cells were used for a total gene expression analysis. Metabolite profiles were measured in plasma using gas chromatography/mass spectrometry. Multivariate statistics was employed to find potential biomarkers for the RA Heat and RA Cold phenotype. A comprehensive biologic interpretation of the results is discussed. Results:The genomics and metabolomics analysis showed statistically relevant different gene expression and metabolite profiles between healthy controls and RA patients as well as between the RA Heat and RA Cold group. Differences were found in the regulation of apoptosis. In the RA Heat group caspase 8 activated apoptosis seems to be stimulated while in the RA Cold group apoptosis seems to be suppressed through the Nrf2 pathway. Conclusions:RA patients could be divided in 2 groups according to CM theory. Molecular differences between the RA Cold and RA Heat groups were found which suggest differences in apoptotic activity. Subgrouping of patients according to CM diagnosis has the potential to provide opportunities for better treatment outcomes by targeting Western or CM treatment to specific groups of patients.

Cold and heat pattern of rheumatoid arthritis in traditional Chinese medicine: distinct molecular signatures indentified by microarray expression profiles in CD4-positive T cell

The research is aimed to explore the distinct molecular signatures in discriminating the rheumatoid arthritis patients with traditional Chinese medicine (TCM) cold pattern and heat pattern. Twenty patients with typical TCM cold pattern and heat pattern were included. Microarray technology was used to reveal gene expression profiles in CD4+ T cells. The signal intensity of each expressed gene was globally normalized using the R statistics program. The ratio of cold pattern to heat pattern in patients with RA at more or less than 1:2 was taken as the differential gene expression criteria. Protein–protein interaction information for these genes from databases was searched, and the highly connected regions were detected by IPCA algorithm. The significant pathways were extracted from these subnetworks by Biological Network Gene Ontology tool. Twenty-nine genes differentially regulated between cold pattern and heat pattern were found. Among them, 7 genes were expressed significantly more in cold pattern. Biological network of protein–protein interaction information for these significant genes were searched and four highly connected regions were detected by IPCA algorithm to infer significant complexes or pathways in the biological network. Particularly, the cold pattern was related to Toll-like receptor signaling pathway. The following related pathways in heat pattern were included: Calcium signaling pathway; cell adhesion molecules; PPAR signaling pathway; fatty acid metabolism. These results suggest that better knowledge of the main biological processes involved at a given pattern in TCM might help to choose the most appropriate treatment.

A network-based analysis of traditional Chinese medicine cold and hot patterns in rheumatoid arthritis

OBJECTIVE Rheumatoid arthritis (RA) is a heterogeneous disease, and traditional Chinese medicine (TCM) can be used to classify RA into different patterns such as cold and hot based on its clinical manifestations. The aim of this study was to investigate potential network-based biomarkers for RA with either a cold or a hot pattern. METHOD Microarray technology was used to reveal gene expression profiles in CD4(+) T cells from 21 RA patients with cold pattern and 12 with hot pattern. A T-test was used to identify significant differences in gene expression among RA patients with either cold or hot pattern. Cytoscape software was used to search the existing literature and databases for protein-protein interaction information for genes of interest that were identified from this analysis. The IPCA algorithm was used to detect highly connected regions for inferring significant complexes or pathways in this protein-protein interaction network. Significant pathways and functions were extracted from these subnetworks by the Biological Network Gene Ontology tool. RESULT Four genes were expressed at higher levels in RA patients with cold pattern than in patients with hot pattern, and 21 genes had lower levels of expression. Protein-protein interaction network analysis for these genes showed that there were four highly connected regions. The most relevant functions and pathways extracted from these subnetwork regions were involved in small G protein signaling pathways, oxidation-reduction in fatty acid metabolism and T cell proliferation. CONCLUSION Complicated network based pathways appear to play a role in the different pattern manifestations in patients with RA, and our results suggest that network-based pathways might be the scientific basis for TCM pattern classification.

Correlation between cold and hot pattern in traditional Chinese medicine and gene expression profiles in rheumatoid arthritis

Clinical manifestations of rheumatoid arthritis (RA) are diversified, and based on the manifestations, the patients with RA could be classified into different patterns under traditional Chinese medicine. These patterns decide the selection of herbal prescription, and thus they can help find a subset of rheumatoid arthritis patients for a type of therapy. In the present study, we combine genome-wide expression analysis with methods of systems biology to identify the functional gene networks for the sets of clinical symptoms that comprise the major information for pattern classification. Clinical manifestations in rheumatoid arthritis were clustered with factor analysis, and two factors (similar to cold and hot patterns in traditional Chinese medicine) were found. Microarray technology was used to reveal gene expression profiles in CD4+ T cells from 21 rheumatoid arthritis patients. Protein-protein interaction information for these genes from databases and literature data was searched. The highly-connected regions were detected to infer significant complexes or pathways in this protein-protein interaction network. The significant pathways and function were extracted from these subnetworks using the Biological Network Gene Ontology tool. The genes significantly related to hot and cold patterns were identified by correlations analysis. MAPK signalling pathway, Wnt signaling pathway, and insulin signaling pathway were found to be related to hot pattern. Purine metabolism was related to both hot and cold patterns. Alanine, aspartate, and tyrosine metabolism were related to cold pattern, and histindine metabolism and lysine degradation were related to hot pattern. The results suggest that cold and hot patterns in traditional Chinese medicine were related to different pathways, and the network analysis might be used for identifying the pattern classification in other diseases.

Anti-oxidative and TNF-α suppressive activities of puerarin derivative (4AC) in RAW264.7 cells and collagen-induced arthritic rats

Puerarin is a major active ingredient extracted from the root of P. lobata, a traditional Chinese herb, and possesses anti-oxidative and anti-inflammatory activities. However, the low oral bioavailability of puerarin limits its further application. Therefore, we synthesized tetraacetyl puerarin (4AC) through acetylation to improve its liposolubility and bioavailability. In the present investigations, we tested the anti-oxidative and TNF-α suppressive activity of 4AC in lipopolysaccharide (LPS)-induced RAW264.7 macrophages and bovine type II collagen-induced arthritic (CIA) rats. The results showed that 4AC retained the bioactivity of puerarin. And 4AC significantly increased the activity of SOD and reduced the level of MDA both in vitro and in vivo. It also improved the level of GSH-PX and the total antioxidant capacity in vivo. Furthermore, it dramatically decreased TNF-α level in the cultured supernatant of RAW264.7 cells treated with LPS and in the serum of CIA rats. These initial results indicated that 4AC had a potential therapeutic effect on CIA rats through an anti-oxidative and TNF-α suppressive activity. In addition, the molecular mechanism of anti-oxidation of 4AC was explored by testing the MAPKs/NF-κB signaling pathway. The results showed that 4AC significantly inhibited NF-κB expression and down-regulated the levels of p-ERK and p-JNK in LPS-activated RAW264.7 cells. These results indicated that 4AC had bioactive anti-oxidative effects and suggest the potential value of 4AC for the treatment of rheumatoid arthritis.

Association between Serum Ferritin Levels and Risk of the Metabolic Syndrome in Chinese Adults: A Population Study

Ferritin is a ubiquitous intracellular protein that can store and release iron and act as a buffer against iron deficiency and iron overload. Ferritin is widely used as a clinical biomarker to evaluate iron status. Increased serum ferritin concentrations have been reported to be associated with metabolic syndrome (MetS) features. However, serum ferritin concentrations differ significantly according to sex and ethnicity, and the data concerning the relationship between serum ferritin concentrations and MetS in Asian men and women are conflicting. This study aimed to explore the relationship between serum ferritin and MetS in Chinese population. Fasting blood samples and anthropometric data collected on 8,441 adults aged 18 and older in 2009 as part of the China Health and Nutrition Survey, a large-scale longitudinal, household-based survey in China. Data was collected by trained physicians and biomarkers were measured with Hitachi Clinical Autoanalyzer 7600 D model and P model. Median levels of serum ferritin were significantly higher in men compared with women (121.9 vs. 51.0 ng/ml, P < 0.001), and significantly lower in non metabolic syndrome population with MetS population (73.2 vs. 106.0 ng/ml, P < 0.001). The difference remained significant after further adjusted for age, nationality, Body mass index (BMI), smoking status, and alcohol consumption. For both men and women, the highest prevalence of MetS occurred in the highest quartile of serum ferritin. The odds ratios increased progressively across the ferritin quartiles (P<0.001 for trend). Increased serum ferritin concentrations are associated with the metabolic syndrome among men and women in China.

The effect of triptolide on CD4+ and CD8+ cells in Peyer's patch of SD rats with collagen induced arthritis.

Triptolide is a purified component from a traditional Chinese herb Tripterygium wilfordii Hook F. It has been shown to have anti-inflammatory and immunosuppressive activities by its inhibitory effect on T cells. But the effect of triptolide on Peyers patch cells is unknown. Enteric mucosal immune system, including Peyers patch, is regarded as one of the sites for inducing immunity tolerance, and this intolerance effect has been used to induce oral tolerance which can considerably reduce arthritis severity in several models of experimental polyarthritis and RA patients. In this study, we investigated the effect of triptolide on the Peyers patch cells and peripheral lymphocytes in collagen induced arthritis (CIA) in rats. CIA in rat is a widely studied animal model of inflammatory polyarthritis with similarities to rheumatoid arthritis (RA). Our data show that triptolide could lower the arthritic scores and delay the onset of CIA. There are more Peyers patches in triptolide treated rats than in control rats, while there is no difference in Peyers patch numbers between CIA rats and triptolide treated rats. In the Peyers patch, more CD4+ cells are observed in CIA rats, and the numbers of CD4+ cells in triptolide treated rats and control rats are similar. While more CD8+ cells are observed in triptolide treated rats, and the numbers of CD8+ cells in CIA rats and control rats are similar. In periphery, more CD4+ cells and less CD4+ cells in CIA rats and triptolide treated rats are respectively observed. Therefore, the regulation on Peyers patch might explain some of the immunosuppressive activities of triptolide, and enteric immune response might be actively involved in CIA pathogenesis. It is suggested that the Peyers patch is one of the primary targets of the immunosuppressive activity of triptolide.

Network-based gene expression biomarkers for cold and heat patterns of rheumatoid arthritis in traditional chinese medicine.

In Traditional Chinese Medicine (TCM), patients with Rheumatoid Arthritis (RA) can be classified into two main patterns: cold-pattern and heat-pattern. This paper identified the network-based gene expression biomarkers for both cold- and heat-patterns of RA. Gene expression profilings of CD4+ T cells from cold-pattern RA patients, heat-pattern RA patients, and healthy volunteers were obtained using microarray. The differentially expressed genes and related networks were explored using DAVID, GeneSpring software, and the protein-protein interactions (PPI) method. EIF4A2, CCNT1, and IL7R, which were related to the up-regulation of cell proliferation and the Jak-STAT cascade, were significant gene biomarkers of the TCM cold pattern of RA. PRKAA1, HSPA8, and LSM6, which were related to fatty acid metabolism and the I-κB kinase/NF-κB cascade, were significant biomarkers of the TCM heat-pattern of RA. The network-based gene expression biomarkers for the TCM cold- and heat-patterns may be helpful for the further stratification of RA patients when deciding on interventions or clinical trials.

Comparison of toxic reaction of Tripterygium wilfordii multiglycoside in normal and adjuvant arthritic rats.

AIM OF THE STUDY Tripterygium wilfordii multiglycoside (GTW), an authorized Chinese patent drug, is used for treatment of rheumatoid arthritis and other immune disease. This study was to determine whether GTW induced different toxic reactions in adjuvant arthritis rats (AA rats) compared to those in normal rats. MATERIALS AND METHODS To prepare arthritic rat model, male Sprague-Dawley (SD) rats were immunized by injecting complete Freunds Adjuvant into right hind footpad. And then, GTW was given to rats intragastrically at dosage of 7 or 105 mg kg(-1)day(-1) from day 15 to day 28 after immunization. Routine clinical parameters and histopathologic changes of liver, kidney and testis were examined. Metabolic profiling in serum of groups was analyzed by LC-MS. A principal component analysis (PCA) and partial-least-squares discriminate analysis (PLS-DA) were carried out combined with mass spectrometry (MS) data set. All the quantitative data were performed by two-way ANOVA analysis following Students t-test. RESULTS AND CONCLUSIONS Treatment with GTW at both doses could diminish the right and left hind paws swelling. There was slight lipoid degeneration in hepatic tissue of normal rats treated by high dose of GTW, but there were not distinctly pathological changes in hepatic tissue of AA rats treated by GTW. Compared normal rats administered with GTW, no statistically significant difference in the serum alanine aminotransferase (ALT), creatinine (CRE), and blood urea nitrogen (BUN) levels were observed. However, the serum aspartate aminotransferase (AST) level was significant decreased in AA rats under exposure GTW compared with normal rats in the same conditions (p<0.05), which indicated that GTW could offer a different liver toxic reaction in normal and AA rats. The metabolic analysis showed that a clear separation of PCA and PLS-DA score spot in normal rats, but not separation was seen in AA rats perturbed with low dosage GTW. The result indicated low dosage GTW might arouse a general toxic in normal rats but not in AA rats. The biomarker analysis showed that the level of lysophosphatidylcholines (LPCs) was down-regulated, but the level of ursodeoxycholic acid (UDCA) and chenodexycholic acid (CDCA) was up-regulated in AA rats compared with normal rats under exposure GTW. According to pathway analysis of metabolic markers, we conceived that LPC, UDCA and CDCA were the critical intermediates of choline and fatty acid metabolism. And the lipid metabolism was a correlative outcome of GTW induced toxicity in the liver in physiological condition animals. Taken together, GTW could induce different toxic reactions between normal and AA rats, and the lipid metabolism might be part of the mechanism for the hepatic lipidosis or the other liver injury.

The effect of triptolide on CD4+ and CD8+ cells in the Peyer's patch of DA rats with collagen induced arthritis.

Triptolide is a purified component from a traditional Chinese herb Tripterygium wilfordii Hook F. It has been shown to have anti-inflammatory and immunosuppressive activities by its inhibitory effect on T-cells. But the effect of triptolide on enteric mucosal immune responses is not well known. The enteric mucosal immune system, especially the Peyers patch, is regarded as one of the sites for inducing immunity tolerance, and this intolerance effect has been used to induce oral tolerance, which can considerably reduce arthritis severity in several models of experimental polyarthritis and rheumatoid arthritis (RA) patients. In this study, we investigated the effect of triptolide on the CD4+ and CD8+ cell distribution in Peyers patch cells and periphery lymphocytes and TGF-β and IFN-γ levels in periphery in collagen induced arthritis (CIA) in DA rats. CIA in the rat is a widely studied animal model of inflammatory polyarthritis, with similarities to RA, and the DA rat is one of most susceptible strains for CIA. Our data show that triptolide could lower the arthritic scores of CIA. The more CD8+ cells in the Peyers patch, the more CD4+ cells in periphery. High level IFN-γ and low level TGF-β in periphery are observed in CIA rats, while the less CD4+ and CD8+ cells in the Peyers patch, the less CD4+ cells in periphery. Low level IFN-γ and high level TGF-β in periphery are shown in triptolide-treated rats. The dose dependency of triptolide was observed in periphery CD4 cells and in the arthritic score. Therefore, the effect of triptolide on Peyers patch immune cells might partially explain some of the immunosuppressive activities of triptolide.