Effects of acupuncture on the number of associated protein phosphorylation in brain tissues of MCAO rats based on protein microarray technique

Objective： To investigate the effects of acupuncture on the number of associated phosphorylated proteins in brain tissues of middle cerebral artery occlusion （MCAO） rats, based on the protein microarray technique. Methods： The MCAO model was prepared according to the modified occlusion method using occlusion lines. Forty healthy Sprague-Dawley （SD） rats were randomly divided into 4 groups using the lottery method： a sham operation group, a model group, a control point group and an acupoint group, with 10 rats in each group. Rats in the sham operation group and the model group only received binding without acupuncture. Rats in the acupoint group received acupuncture at Dazhui （GV 14）, Baihui （GV 20） and Shuigou （GV 25）; rats in the control point group received acupuncture at non-acupoint control points. The needle was twisted once for 1 min after insertion and another time in the middle of the 30 min needle retaining. Acupuncture was conducted once every 12 h for 6 consecutive times. At the end of the experiment, the neurological impairment score was collected, and cells of the ischemic brain tissues were extracted. The protein phosphorylation of the related signaling was detected using the 720 phosphorylated antibody microarray technique, and the differentially expressed proteins between groups were screened. Results： The neurological impairment scores after 72 h of treatment： compared with the sham operation group, the scores of the model group, the control point group and the acupoint group were significantly increased （P〈0.01）; compared with the model group, the scores of the acupoint group and the control point group were significantly decreased （P〈0.01,P〈0.05）; the score of the acupoint group was better than that of the control point group （P〈0.05）. The results of the protein microarray： compared with the sham operation group, 48 proteins showed up-regulated phosphorylation （≥1.5 times） in the model group and the down-regulated was 28; compared with the model group, 35 proteins showed up-regulated phosphorylation in the control point group, and the down-regulated was 24. There were 29 proteins showing up-regulated phosphorylation in the acupoint group and the down-regulated was 51. The numbers of proteins involved in the function and signal transduction pathways were also different. Conclusion： Acupuncture at Dazhui （GV 14）, Baihui （GV 20） and Shuigou （GV 25） can effectively repair brain injury. The ischemic injury of brain tissue may be caused by imbalance of a variety of proteins, and acupuncture can promote brain tissue repair by multi-functional and multi-channel regulation of the protein disorders.

Applied research on serum protein fingerprints for prediction of Qi deficiency syndrome and phlegm and blood stasis in patients with non-small cell lung cancer

OBJECTIVE:This study screened serum tumor biomarkers by surface enhanced laser desorption/ionization time-of-flight mass spectrometry(SELDI-TOF-MS) to establish a subset which could be used for the prediction of Qi deficiency syndrome and phlegm and blood stasis in patients with non-small cell lung cancer;and as diagnostic model of Chinese medicine.METHODS:Serum samples from 63 lung cancer patients with Qi deficiency syndrome and phlegm and blood stasis,and 28 lung cancer patients with non-Qi deficiency syndrome and phlegm and blood stasis were analyzed using SELDI-TOF-MS with a PBS II-C protein chip reader.Protein profiles were generated using immobilized metal affinity capture(IMAC3) protein chips.Differentially-expressed proteins were screened.Protein peak clustering and classification analyses were performed using Biomarker Wizard and Biomarker Pattern software packages,respectively.RESULTS:A total of 268 effective protein peaks were detected in the 1,000-10,000 Da molecular range for the 15 serum proteins screened(P<0.05).The decision tree model was M 2284.97,with a sensitivity of 96.2% and a specificity of 66.7%.CONCLUSION:SELDI-TOF-MS techniques,combined with a decision tree model,can help identify serum proteomic biomarkers related to Qi deficiency syndrome and phlegm and blood stasis in lung cancer patients;and the predictive model can be used to discriminate between Chinese medicine diagnostic models of disease.

Influence of irbesartan on the urinary excretion of cytokines in patients with chronic kidney disease

Background The non-hemodynamic effects of angiotensin receptor blocker （ARB） in the delay of progression of chronic kidney disease （CKD） remain unclear. In this study, we investigated the influence of irbesartan on the urinary excretion of cytokines in patients with CKD. Methods In this randomized perspective clinical trial, different doses of irbesartan （150 mg/d and 300 mg/d） were given to two groups of patients in a cross-over design. Blood pressure （BP）, creatinine clearance （Ccr） and 24-hour proteinuria were examined. Urinary excretion of cytokines was determined by human inflammatory cytokine antibody array. A two-fold change in spot intensity was considered significant. Results Urinary excretion of cytokines （granulocyte colony stimulating factor （GCSF）, intercellular cell adhesion molecule-1 （ICAM-1）, interferon y （IFN-y）, interleukin 1β （IL-1β）, IL-2, IL-6, IL-8, IL-11, IL-15 and macrophage inflammatory protein 1δ （MIP-Iδ） in group B （irbesartan 300 mg/d） was significantly decreased in comparison to group A （irbesartan 150 mg/d） after 8-week treatment. In group A, 8 weeks of treatment induced a two- to nine-fold reduction in urinary cytokine levels （GCSF, GM-CSF, IFN-γ, IL-1α, IL-11, IL-12p40, MCP-2, MIP-1α）, while increasing the dosage to 300 mg/d further decreased the excretion of GCSF, GM-CSF, IL-12p40, MCP-2 and MIP-1α by week 18. There was no significant difference in BP or Ccr between the two groups. However, 24-hour proteinuria was significantly reduced in both groups, and in group A the reduction was dose dependent.Conclusion Irbesartan offers additional renoprotection in a dose-dependent manner by reducing pro-inflammatory cvtokines excretion in the urine of CKD patients.

Apoptosis of non-tumor cells contributes to increased serum cytochrome c level in a neuroblastoma xenograft model

Background Neuroblastoma （NB） is one of the most common malignant solid tumors of childhood.It is still not clear whether the apoptosis of tumor cells or the non-tumor cells contributes to the increase of concentration of cytochrome c （Cyt c） in the serum of the cancer patients.The aim of this research was to identify the source of the Cyt c in the serum when the tumor grows up by subcutaneous inoculation of human NB cells into nude mice.Methods We subcutaneously inoculated human NB cells （KP-N-NS） into nude mice and collected the sera of tumor-bearing mice （n=14） and control mice （n=25） 4 weeks later in order to screen for and identify differentially expressed proteins in the serum.Differentially expressed proteins in the serum were screened by surface-enhanced laser desorption/ionization-time-of-fiight （SELDI-TOF） mass spectrometry.Results The relative intensity of a protein having a mass-to-charge ratio （m/z） of 11 609 was 3338.37±3410.85 in the tumor group and 59.84±40.74 in the control group,indicating that the expression level of this protein in the tumor group was 55.8 times higher than that in the control group.Serum proteins were separated and purified by high-performance liquid chromatography （HPLC）.Liquid chromatography coupled with tandem mass spectrometry （LC-MS/MS） was performed to produce peptide mass fingerprints （PMFs）.Spectrum analysis and a database search revealed that the highly expressed protein （m/z=11605.4） from the serum of tumor-bearing mice was the mouse Cyt c.Conclusions Increased concentration of Cyt c in the serum of tumor-bearing nude mice might be partially attributed to the secretion of this protein by non-tumor cells.