Berberine protects endothelial progenitor cell from damage of TNF-α via the PI3K/AKT/eNOS signaling pathway.

Endothelial progenitor cells (EPCs) dysfunction is closely correlated with the coronary artery injury induced by Kawasaki disease (KD). The level of tumor necrosis factor-α (TNF-α) elevated significantly in acute phase of KD which can damage the functions of EPCs. The aim of this study was to investigate whether berberine (BBR) can protect EPCs from the inhibition caused by TNF-α via the PI3K (Phosphatidyl Inositol 3-kinase) /AKT (Serine/threonine protein kinase B) /eNOS (endothelial Nitric Oxide synthase) signaling pathway. The cell proliferative ability of EPCs was determined by MTT (methyl thiazolyl tetrazolium) assays. Nitric oxide (NO) level was determined in supernatants. The mRNA level of eNOS, PI3K and AKT were measured by Real Time-Polymerase Chain Reaction (RT-PCR), and the protein levels of eNOS, phospho-eNOS (p-eNOS), Akt, phospho-Akt (p-Akt) and PI3K were analyzed using Western-blot. The results demonstrated that TNF-α inhibits the proliferative ability of EPCs. However, BBR improves the proliferative activity of EPCs inhibited by TNF-α. Blockade of PI3K by 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (Ly294002) and blockade of eNOS by l-NAME (NG-Nitroarginine Methyl Ester) attenuates the effect of BBR. BBR can increase the level of PI3K/Akt/eNOS mRNA and the protein level of PI3K, p-Akt, eNOS and p-eNOS, which can be blocked by PI3K inhibitor (LY294002) and eNOS inhibitor (l-NAME). Therefore, we concluded that impaired EPCs proliferation could be reversed by BBR via the PI3K/AKT/eNOS signaling pathway.

[Effects of intravenous immunoglobulin and aspirin treatment on the functions of circulating endothelial progenitor cells in children with Kawasaki disease].

OBJECTIVE: To study the effects of intravenous immunoglobulin (IVIG) and aspirin treatment on the functions of circulating endothelial progenitor cells (EPCs) in children with Kawasaki disease (KD) and possible mechanisms. METHODS: Blood samples were obtained in 10 children with KD before and 7 days after the treatment by IVIG and aspirin. MTT method, modified Boyden chamber method and cell culture plate adhesion method were used to assess the functions of EPCs, including proliferation, adhension and migration activities. The plasma levels of tumor necrosis factor-α (TNF-α) and high-sensitivity C reactive protein (hs-CRP) were also measured. RESULTS: The functions of circulating EPCs 7 days after IVIG and aspirin treatment were significantly improved. IVIG and aspirin treatment significantly reduced plasma TNF-α and hs-CRP concentrations. There was a significant linear regression relationship between the reduced plasma TNF-α and hs-CRP levels and the increased functions of circulating EPCs. CONCLUSIONS: IVIG and aspirin treatment can improve the functions of circulating EPCs, possibly through reducing plasma concentrations of TNF-α and hs-CRP.

The functions of endothelial progenitor cells were significantly improved after treatment with intravenous immunoglobulin and aspirin in children with Kawasaki disease.

We sought to determine the effects of treatment with intravenous immunoglobulin (IVIG) and aspirin on the functions of endothelial progenitor cells (EPCs) in patients with Kawasaki disease (KD) as well as its relationship with concentrations of tumor necrosis factor-α (TNF-α) and high-sensitivity C-reactive protein (hs-CRP). Ten KD patients in the acute phase of their disease were recruited. We investigated EPC functions in children with KD before and after treatment with IVIG and aspirin. In vitro assays were used to measure the functions, including proliferation, adhesion, and migration activities, of EPCs. Plasma levels of TNF-α and hs-CRP were also assessed. All of the data were assessed before and at 7 days after treatment initiation. EPC functions after 7 days of treatment with IVIG and aspirin were significantly improved than they were before treatment with IVIG and aspirin. Treatment with IVIG and aspirin significantly decreased TNF-α and hs-CRP concentrations. There was a significant linear regression relationship between decreased plasma TNF-α levels, hs-CRP levels, and increased functions of circulating EPCs. The results of our study indicate that the functions of circulating EPCs improved after treatment with IVIG and aspirin, which may be related to decreased concentrations of TNF-α and hs-CRP.

[Decreased circulating endothelial progenitor cell function: relationship with serum concentrations of high-sensitivity C-reactive protein in children with Kawasaki disease].

OBJECTIVE: To study the function of circulating endothelial progenitor cells and its relationship with serum concentrations of high-sensitivity C-reactive protein (Hs-CRP) in children with Kawasaki disease. METHODS: Ten children with Kawasaki disease and ten healthy children as a control group were enrolled. The peripheral mononuclear cells were induced into endothelial progenitor cells using Dulbecco's Modified Eagle Medium containing vascular endothelial growth factor and basic fibroblast growth factor. The proliferative ability, migratory ability and adhesive ability of endothelial progenitor cells were assessed by MTT methods, modified Boyden chamber methods and cell culture plate adhesion method, respectively. The concentrations of serum Hs-CRP were measured by latex enhanced turbidimetric immunoassay. RESULTS: The proliferative ability, migratory ability and adhesive ability of endothelial progenitor cells in the Kawasaki disease group were significantly lower than those in the control group (P<0.01). The serum concentrations of Hs-CRP in the Kawasaki disease group were significantly higher than those in the control group (87.1+/-30.2 mg/L vs 5.3+/-3.4 mg/L; P<0.01). The function of circulating endothelial progenitor cells was negatively correlated with serum concentrations of Hs-CRP in the Kawasaki disease group. CONCLUSIONS: The function of circulating endothelial progenitor cells is decreased in children with Kawasaki disease, which may be associated with the abnormal expression of inflammatory mediators.

The number and function of circulating endothelial progenitor cells in patients with Kawasaki disease.

Kawasaki disease (KD) is associated with coronary artery injury. Studies have shown that the endothelial progenitor cell (EPC) participates in the process of arterial repair. Data have been reported that the number of EPC increased significantly in the subacute phase of KD. However, until now, there are no data about the functions of EPC in KD patients. The present study was designed to further investigate the number and functions of EPC in KD. Ten KD patients in the acute phase and ten healthy volunteers were recruited and attributed to the KD group and control group, respectively. The circulating CD34/kinase insert domain-containing receptor double positive cells were evaluated in the two groups using flow cytometry. In vitro assays were used to measure the functions of EPC, including proliferation, adhesion, and migration activities. The plasma levels of nitric oxide (NO), tumor necrosis factor-alpha (TNF-alpha), and high sensitivity C-reactive protein (hs-CRP) were also assessed in both groups. The number of EPC in the KD group was significantly higher than that of the control group (0.021 +/- 0.007% vs. 0.014 +/- 0.003%, P < 0.05). The migratory response of EPC was significantly decreased in the KD group, compared with that of the control group (5.50 +/- 1.78 vs. 3.40 +/- 1.35 cells/high power field, P < 0.01). Similarly, the proliferative and adhesive activities of EPC in the KD group were also decreased (0.47 +/- 0.08 vs. 0.66 +/- 0.07, P < 0.01; 6.5 +/- 2.12 vs. 11.2 +/- 2.04 cells/high power field, P < 0.01). The plasma NO, TNF-alpha, and hs-CRP levels in the KD group were higher than those of the control group (54.10 +/- 11.78 vs. 38.80 +/- 11.10 mumol/l, P < 0.01; 48.20 +/- 7.42 vs. 37.00 +/- 11.12 pg/ml, P < 0.05; 87.10 +/- 30.18 vs. 5.30 +/- 3.37 mg/l, P < 0.01). The number of circulating EPC positively correlated with the level of NO (r = 0.92, P < 0.001), and the functions of EPC negatively correlated with the levels of TNF-alpha and hs-CRP, respectively. In Kawasaki disease, the number of EPC was enhanced and the functions of EPC were attenuated. The two-way regulation of circulating EPC in KD patients may be associated with the disorders of cytokines or messengers in KD patients.

[Effects of grassland restoration from cropland on soil macrofaunal community in Wuchuan County, Inner Mongolia].

By the method of hand-sorting, and using the indices individual number, group number, and biodiversity, the changes in the soil macrofaunal community under the grassland restoration from cropland in Wuchuan County, Inner Mongolia were investigated. A total of 1479 soil macrofauna belonging to 3 classes, 7 orders and 41 families were captured. After the grassland restoration from cropland, the group number, individual number, and biomass of soil macrofauna all had an increasing trend, and the biodiversity of soil macrofaunal community reflected by D(Ma) and DG increased significantly. However, due to the short term of cropland rehabilitation, the Shannon-Wiener index H' and the Pielou evenness index E of the soil microfaunal community in grassland were not higher than those in cropland. Different soil macrofaunal groups had different responses to the grassland restoration from cropland. The preferable approaches for the grassland restoration from cropland were planting Medicago sativa or planting Medicago sativa + Artemisia sp.

[Prognostic factors related to recurrence after extended thymectomy in patients with myasthenia gravis].

OBJECTIVE: To identify the recurrence-related factors in patients with myasthenia gravis (MG) after extended thymectomy. METHODS: Followed up was conducted on 271 MG patients, 127 males and 146 females, aged 31 (4-57), who underwent extended thymectomy for 18-84 months. Post-operational pathological examination showed 32 cases of thymoma and 239 cases of diseases other than thymoma. After operation the patients were treated with pyridostigmine only or combined with adrenocortical hormone. The relevant factors of the 135 patients with relapse were evaluated: sex, Osserman classification, age while being operated on, duration of preoperative period, pathologic type of thymus, use of steroid before operation, infection after operation, whether only taking anticholinesterase drugs after operation, use of steroid immediately after operation, stopping medicine or decreasing the dose of medicine within 1-3 months after remission of symptoms. RESULTS: COX univariate analysis revealed that failure to take steroid immediately after operation (OR = 2.914, P = 0.000), infection after operation (OR = 3.441, P = 0.000), only taking anticholinesterase drugs after operation (OR = 5.947, P = 0.000), and immediately stopping medicine use or decreasing the dose of medicine within 1-3 months after the remission of symptoms (OR = 2.242, P = 0.000) were prognostic factors for postoperative recurrence. On the other hand, multivariate logistic regression analysis revealed that infection after operation (OR = 47.63, P = 0.000), only taking anticholinesterase drugs after operation (OR = 62.38, P = 0.000), and stopping medicine or decreasing the dose of medicine 1-3 months after remission of symptoms (OR = 32.76, P = 0.000) were independent influencing factors of recurrence after operation. CONCLUSION: Post-operative infection, only taking pyridostigmine, and stopping medicine too early are independent factors of postoperative relapse. Regular treatment and timely use of adrenocortical hormone decrease the recurrence after operation.

Dysphonia as a primary manifestation in myasthenia gravis (MG): a retrospective review of 7 cases among 1520 MG patients.

Myasthenia gravis (MG) is an autoimmune disorder of neuromuscular transmission, usually recognized with ocular complaints or generalized muscle weakness. However, among the 1520 MG cases that had been diagnosed and treated in our hospital in the last 15 years (1990-2005), we have identified 7 MG patients whose initial and prominent complaint was dysphonia and all had been misdiagnosed elsewhere. The diagnoses were confirmed with fibrolaryngoscope and voice analysis employed before and after a positive neostigmine (anticholinesterase) test. Electromyography with repetitive stimulations, single-fiber electromyography, and laboratory and radiographic evaluations were also conducted for diagnosis. A surprisingly low seropositivity rate of anti-acetylcholine-receptor antibodies (1/7) and anti-MuSK (Muscle Specific Kinase) antibodies (0/6) were found in these dysphonia MG patients. A cholinesterase inhibitor (ChEI) and immunosuppressive therapy were applied for treatment. Extended thymectomy was applied to MG patients with thymus hyperplasia or thymic tumor. Significant improvement was found in all 7 cases after these treatments. We have developed a sere of diagnostic protocol for this rare type of laryngeal MG, and discussed the clinical implication of our data. In summary, dysphonia or laryngeal disorder can be the only prominent manifestation of MG in rare cases, which should be taken into consideration during the diagnosis to patients with exclusive laryngeal complaints.

[Prognostic factors of myasthenic crisis after extended thymectomy in patients with generalized myasthenia gravis].

OBJECTIVE: To investigate the prognostic factors of myasthenic crisis after extended thymectomy in patients with generalized myasthenia gravis (MG). METHODS: Extended thymectomy was performed on 176 patients with generalized MG, 74 males and 102 females, aged 4 - 67, of which 36 experienced postoperative myasthenic crisis and required prolonged mechanical ventilation. The relations among the age, sex, preoperative course of disease, pathologic type of thymus, Osserman classification, history of infection during 1 month preoperatively, history of myasthenic crisis 1 month preoperatively, thymoma, preoperative daily dose of pyridostigmine, preoperative steroid use, operation time, intra-operative blood loss, and intra-operative pleura injury and postoperative myasthenia crisis were analyzed. RESULTS: Univariate analysis showed that bulbar symptoms (OR = 8.494, P = 0.001), history of myasthenic crisis 1 month preoperatively (OR = 5.667, P = 0.000), thymoma (OR = 2.147, P = 0.047), Osserman types III and IV (OR = 0.459, P = 0.000), history of infection during 1 month preoperatively (OR = 3.30, P = 0.038), large pre-operative dose of pyridostigmine (OR = 1.019, P = 0.001), long operation time (OR = 1.012, P = 0.034), and more blood loss (186 ml +/- 163 ml) (OR = 1.004, P = 0.012), were all prognostic factors or postoperative myasthenic crisis. However, multivariate logistic regression analysis revealed that preoperative bulbar symptoms (OR = 7.709, P = 0.003), history of infection during 1 month preoperatively (OR = 4.582, P = 0.037), history of myasthenic crisis 1 month preoperatively (OR = 4.526, P = 0.001, large pre-operative dose of pyridostigmine (OR = 1.016, P = 0.001) were prognostic factors of postoperative myasthenic crisis. CONCLUSION: Preoperative bulbar symptoms, history of preoperative myasthenic crisis, history of preoperative infection, and large preoperative dose of, pyridostigmine are all independent influencing factors of postoperative myasthenic crisis. Ample preoperative care may prevent postoperative myasthenic crisis in the patients with such factors.

[Long-term efficacy of enlarged thymectomy in treatment of myasthenia gravis and relevant influencing factors: study of 410 cases].

OBJECTIVE: To investigate the long-term efficacy of enlarged thymectomy in treatment of myasthenia gravis (MG) and relevant influencing factors. METHODS: 546 patients with MG underwent enlarged thymectomy and were followed up for 28 months (6 to 85 months). Effective follow-up data were obtained from 410 out of the 546 patients. The clinical data of these 410 patients, 199 males and 211 females, were analyzed. RESULTS: The remission rate was 42.9% and the effective rate was 82.3%. Multivariate logistic regression analysis revealed that short duration of preoperative period was an independent factor of the surgical curative effect (odds ratio = 0.310, P = 0.006) and sex, age, Osserman classification, pathologic type of thymus seemed to be irrelevant to the surgical curative effect. CONCLUSION: Thymectomy is an effective measure for MG and shows a better prognosis in the patients with shorter illness duration.

[Clinical value of the immune parameters in patients with myasthenia gravis].

OBJECTIVE: To evaluate the clinical value of the indices, including IgG, IgA, IgM, CD3, CD4 and CD8. METHODS: Detect CD3, CD4, CD8 by flow cytometry and detect IgG, IgA, IgM by turbidimetric method. We analyzed clinical features according to different classification of immune index, and changes of the level of these immune indices before and after treatment. RESULT: The level of IgG, IgA in patients with generalized MG was higher than that of ocular MG (P < 0.05). The ratio of CD4 to CD8 was classified into 3 groups: low (118 cases), high (60 cases), normal (68 cases). Likewise, the level of IgG, IgA, IgM was classified into 4 groups: low (156 cases), high (65 cases), disorder (31 cases), normal (67 cases). There was no statistically significance on changes of level of these immune indices before and after treatment. There was no statistically significance on clinical materials among different groups. In the low group of humoral immunity, the remission ratio of the patients with the treatment of gamma globulin was higher than that of patients without gamma globulin (95.2%, 28.8%, respectively. P < 0.01). CONCLUSION: The immune indices of IgG, IgA, IgM, CD3, CD4, CD8 seemed to be irrelevant to severity of disease and prognosis. To some extent, these immune indices can be reference in the aspect of treatment.