This study investigated the effects and action mechanism of tannins from Galla chinensis cream(TGCC) on the skin wound of rat tail. Male Sprague Dawley(SD) rats were randomly divided into a control group, model group, model+low-dose TGCC(50 mg per rat) group, model+high-dose TGCC group(100 mg per rat), and model+TGC+FAK inhibitor(Y15) cream(100 mg+10 mg per rat) group, with 10 rats in each group. After the rat tail skin injury model was successfully constructed, in the treatment group, corresponding drugs were applied to the wound surface, while in the control and model groups, the same amount of cream base as the TGCC group was applied by the same method. Then, sterile gauze was wrapped around the wound edge, and these operations were performed three times a day for 28 consecutive days. The wound healing status at the third, seventh, eleventh, fourteenth, twenty-first, and twenty-eighth days was recorded, and the wound healing rate and healing time were calculated. On the day after the last dose of medication, rat serum and tail skin wound tissue were collected for analyzing the activities of serum alanine aminotransferase(ALT), aspartate aminotransferase(AST), creatinine(CREA), urea, reactive oxygen species(ROS), interferon gamma(IFN-γ), interleukin(IL)-1β, IL-6, IL-4, IL-10, tumor necrosis factor(TNF)-α, as well as catalase(CAT), glutathione(GSH), lactate dehydrogenase(LDH), malondialdehyde(MDA), myeloperoxidase(MPO), superoxide dismutase(SOD), total antioxidant capacity(T-AOC), platelet endothelial cell adhesion molecule-1(CD31), and leukocyte differentiation antigen 34(CD34) in the wound tissue of rat tail skin. Hematoxylin-eosin, Masson, and sirius red staining were used to observe the morphological changes in the wound tissue of rat tail skin. The thickness of the epidermis, the number of fibroblasts and blood vessels, and the contents of collagen fibers, typeⅠ collagen(COLⅠ), and COLⅢ were calculated. The mRNA expressions of keratin 10(KRT10), KRT14, vascular endothelial growth factor(VEGF), fibroblast growth factor(FGF), epidermal growth factor(EGF), CD31, CD34, matrix metallopeptidase-2(MMP-2), MMP-9, COLⅠ, COLⅢ, desmin, fibroblast specific protein 1(FSP1), IFN-γ, IL-1β, TNF-α, IL-4, IL-6, and IL-10 in skin wound tissue were determined by quantitative real-time polymerase chain reaction(PCR). Western blot was utilized to detect the protein expressions of KRT10, KRT14, VEGF, FGF, EGF, MMP-2, MMP-9, COLⅠ, COLⅢ, desmin, FSP1, focal adhesion kinase(FAK), phosphorylated focal adhesion kinase(p-FAK), phosphatidylin-ositol-3-kinase(PI3K), phosphorylated phosphatidylin-ositol-3-kinase(p-PI3K), protein kinase B(Akt), phosphorylated protein kinase B(p-Akt), mammalian target of rapamycin(mTOR), and phosphorylated mammalian target of rapamycin(p-mTOR). The results manifest that TGCC can dramatically elevate the healing rate of rat tail wounds and shorten wound healing time. Besides, it can reduce serum ROS levels, the contents of MDA, MPO, and LDH in the rat skin wound tissue, as well as the serum IFN-γ, IL-1β, IL-6, and TNF-α levels and the mRNA expression levels of IFN-γ, IL-1β, IL-6, and TNF-α in the skin wound tissue. It can elevate the activities of CAT, GSH, SOD, and T-AOC in wound tissue, the IL-4 and IL-10 contents in serum, and the mRNA expressions of IL-4 and IL-10 in the wound tissue. In addition, TGGC can inhibit inflammatory cell infiltration and increase the epidermal thickness, counts of fibroblasts and blood vessels, and contents of collagen fibers, COLⅠ, and COLⅢ. Besides, TGCC can elevate the mRNA and protein expressions of epidermal differentiation markers(KRT10 and KRT14), endothelial cell markers(CD31 and CD34), angiogenesis and fibroblast proliferation, differentiation markers(VEGF, FGF, EGF, COLⅠ, COLⅢ, desmin, and FSP1), reduce the mRNA and protein expressions of gelatinases(MMP-2 and MMP-9), and increase protein expressions of p-FAK, p-PI3K, p-Akt, p-mTOR, as well as ratios of p-FAK/FAK, p-PI3K/PI3K, p-Akt/Akt, and p-mTOR/mTOR. These results suggest that TGCC can significantly facilitate skin wound healing, and its mechanism may be related to the activation of the FAK/PI3K/Akt/mTOR signaling pathway, inhibition of inflammatory cell infiltration in skin wound tissue, elevation of epidermal thickness, counts of fibroblasts and vessels, and contents of collagen fiber, COLⅠ, and COLⅢ, and reduction of MMP-2 and MMP-9 expressions, thus accelerating wound healing.
Animals ; Male ; Wound Healing/drug effects* ; Rats ; Rats, Sprague-Dawley ; Signal Transduction/drug effects* ; TOR Serine-Threonine Kinases/genetics* ; Phosphatidylinositol 3-Kinases/genetics* ; Skin/metabolism* ; Proto-Oncogene Proteins c-akt/genetics* ; Tannins/pharmacology* ; Humans ; Drugs, Chinese Herbal/administration & dosage* ; Focal Adhesion Kinase 1/genetics*

OBJECTIVE To establish a method for simultan eous determination of 5 kinds of pentacyclic triterpenoids as 3-O- acetyl-pomolic acid in Chaenomeles speciosa ，and to analyze the difference in the contents of C. speciosa from different producing areas by different processing technologies . METHODS HPLC method was adopted . The determination was performed on COSMOSIL 5 C18-MS-Ⅱ column with mobile phase consisted of acetonitrile -0.005 mol/L ammonium dihydrogen phosphate solution （pH value adjusted to 6.5 with phosphoric acid ）（70∶30，V/V）at the flow rate of 1.0 mL/min. The column temperature was set at 30 ℃. The detection wavelength was set at 210 nm，and sample size was 20 μL. The contents of the other four pentacyclic triterpenoids were calculated according to quantitative analysis of multi -components by single -marker（QAMS）using oleanolic acid as internal reference . The results were compared with those determined by external standard method . The total content of oleanolic acid and ursolic acid ，the total content of 5 components in C. speciosa from different producing areas and different processing technologies were compared . RESULTS The linear range of 3-O-acetyl-pomolic acid ，betulinic acid ，oleanolic acid ，ursolic acid and 3-O-acetyl ursolic acid were 4.06-81.2，2.12-42.4，9.62-192.3，7.77-155.4，4.21-84.1 μg/mL，respectively（R2＞0.999）. RSDs of precision ，reproducibility and stability （24 h）tests were all lower than 3%. The average recoveries were 98.29%-101.38% （RSDs＜3%，n＝6）. The mass fraction of 3-O-acetyl-pomolic acid ，betulinic acid ，ursolic acid and 3-O-acetyl ursolic acid measured by QAMS w ere 0.023%-0.060%，0.044%-0.528%，0.101%-0.368%，0.067%-0.221%，respectively；the deviations from the results measured by external standard method were all within 8%. The total content of oleanolic acid and ursolic acid， the total content of 5 components in C. speciosa processed by fresh -cut technology from the same producing area were higher than those in C. speciosa processed by traditional technology ，and the total content of 5 components in C. speciosa from Chongqing Qianjiang were significantly higher than those from other producing areas （P＜0.05）. CONCLUSIONS QAMS method is established for the simultaneous determination of 3-O-acetyl-pomolic acid ，betulinic acid ，oleanolic acid ，ursolic acid and 3-O-acetyl ursolic acid in C. speciosa. Established method is simple ，rapid and accurate . The total content of 5 components in C. speciosa produced in Chongqing Qianjiang is higher ，and the total content of C. speciosa processed by fresh -cut technology from the same origin is higher than C. speciosa processed by traditional technology .

Antimicrobial resistance is on the rise while the number of antibiotics being brought to market continues to drop. Drug-resistant genes and drug-resistant bacteria infection have seriously threatened human health. Therefore, antimicrobial resistance presents an ongoing challenge that requires multifaceted approaches including: biomedical innovation; improved surveillance of antibiotic consumption and antimicrobial resistance generated rates; prevention of health-care-associated infections and transmission of multidrug-resistant bacteria and environmental dissemination; rapid microbiological diagnosis; and curtailed clinical and veterinary misuse. Fortunately, combating antimicrobial resistance has been highly valued and supported by the government, scientists and entrepreneurs of various countries. With the continuous introduction of new technologies, new products, and new management measures, the problem of antimicrobial resistance must be controlled and alleviated.

Objective To investigate the secretory capacity and apoptosis of interleukin ( IL)-21 induced normal B cells by co-culture with serum from patients with systemic lupus erythematosus (SLE). Methods Serum from twenty new-onset SLE patients and 20 healthy donors were collected .CD1+9 B cells from the normal controls were co-cultured with serum from SLE patients in the presence or absence of IL-21-R-FC(4 μg/ml).Supernatant IgG and IgM concentration were measured by immunoturbidimetric assay on day 5.Supernatant anti-dsDNA level was determined by ELISA .The percentage of apoptotic cells was detected by flow cytometer.Results IgG,IgM and anti-dsDNA levels in normal B cells with SLE serum were significantly higher than those in the serum of SLE patients alone [ ( 5.84 ±1.79 ) g/L vs ( 4.25 ± 1.48)g/L,P=0.000;(0.46 ±0.21)g/L vs (0.43 ±0.21)g/L,P=0.003;(127.76 ±70.24)IU/ml vs (115.15 ±63.88) IU/ml,P=0.014 respectively].However, no significant differences were found in the group of normal B cells with non-homologous serum from normal controls (P>0.05).Supernatant IgG, IgM and anti-dsDNA levels in normal B cells with SLE serum significantly decreased while IL-21R-fusion protein was added [(5.26 ±1.62)g/L vs (5.84 ±1.79)g/L, P=0.006;(0.42 ±0.20)g/L vs (0.46 ±0.21)g/L, P=0.002;( 118.00 ±69.62 ) IU/ml vs ( 127.76 ±70.24 ) IU/ml, P =0.012 respectively ] .The apoptotic rate of B cells with SLE serum was significantly higher than that with normal serum [ ( 47.88 ± 12.65)%vs (38.86 ±10.32)%,P =0.004].But adding IL-21R-fusion conversed the apoptotic rates [(42.08 ±12.52)%vs (47.88 ±12.65)%,P=0.001].Conclusions SLE serum could induce normal B cells to form immunoglobulin secreting cells and producing autoantibodies , or apoptosis in pathological conditions.IL-21 might be considered as a potential therapeutic target of SLE .

Objective To obtain the soluble single-chain fragment V (ScFv)monoclonal antibodies (McAbs) against the SSA antigen epitopes.Methods Three octapeptides (60 000 SSA antigen residues 482-493 termed as P1 epitope, residues 310-323 termed as P2 epitope and residues 230-241 termed as P3 epitope) were synthesized on the lysine frame.The McAbs were panned by coating the corresponding as targets.The specificity, affinity and gene squences of the positive clones were assessed.Soluble single-chain fragment V antibodies special for SSA antigen epitopes were expressed and then identificated.Results After 5 rounds of panning, reactive scFv clones contained full-length scFv antibodies coding regions were obtained,with sufficient affinity and specificity for respective antigen peptides.The absorbance values at 410 nm of the fusion protein of anti-P1-P3 activity with the corresponding peptides were 1.43 ± 0.23, 0.82 ±0.31 and 0.80 ± 0.25, and there was also statistically significant difference in the cross reactions ( P ＜ 0.01 ).Three clones were successfully expressed and then purified by His-bind resin.The activity in vivo of soluble ScFv antibodies was identified to be positive by the indirect immune-fluorescence assay on Hep-2 cells.Conclusion Souble ScFv McAbs against corresponding SSA antigen peptides with high affinity,specificity and activity in vivo were obtained, which are to be competent enough for epitopes expression on the target organs.

Objective To compare the T cell receptor recombination excision cycle (TREC) levels in peripheral blood mononuclear cells (PBMC) of systemic lupus erythematosus (SLE) patients with normal age- and gender- matched controls. To investigate the correlations between TREC levels of SLE patients and their clinical features. Methods We studied TREC levels in peripheral blood mononuclear cells (PBMC) of 21 SLE patients and 22 normal age- and sex- matched controls. TREC concentration was determined by real-time quantitative polymerase chain reaction (real-time qPCR) as the number of TREC copies/1000 PBMCs. The clinical features of the SLE patients such as systemic lupus erythematosus disease activity index (SLEDAI) , ESR, C reaction protein (CRP) , ANA, anti-dsDNA and complement levels and organ involvement were recorded and assessed. Results SLE patients had lower TREC levels [ (9.6 ± 7.5 )copies/1000 PBMC] than controls[ (16.1 ±11.1) copies/1000 PBMC,P = 0.033]. There was an inverse correlation between age and TREC levels in controls (r =- 0. 614, P = 0. 002) but not in SLE patients.There was an inverse correlation between SLEDAI and TREC levels in SLE patients(r =-0. 656, P =0. 001) and TREC levels seemed to have relations to skin lesions ( r = - 0. 620, P = 0. 003 ). No other clinical association was observed between TREC levels and clinical and laboratory SLE manifestations.Conclusion SLE patients had lower TREC levels than normal controls and there is a tendency that TREC level is reversely correlated with disease activity. The decrease PBMC TREC level is indicative of a low proportion of recent thymic emigrant (RTE) in SLE and could be caused by decreased RTE output and/or by increased peripheral T cell proliferation in this disease. The under-representation of RTE in the peripheral T cell pool may play a role in the immune tolerance abnormalities observed in SLE.

Objective To analize the clinical features of patients with hemophagocytic syndrome (HI'S). Methods The underlying diseases, clinical characteristics, laboratory findings, responses to therapy as well as their outcomes of 45 patients with HPS were analyzed retrospectively. Results The underlying diseases included virus infection (n = 9) , tuberculosis (n =2), rheumatic fever (n =4) , malignancies (n =22) , unknown diseases (n =8). HPS was clinically characterized by high fever(93. 3% ) , hepatomegaly (77. 8% ) , splenomegaly(80% ) , lymphadenopathy (55. 6% ) , skin rash(24.4% ), gastrointestinal hemorrhage(22. 2% ) , renal (35. 6% ) and central nervous systern involvement( 15. 6% ) , five patients presented with disseminated intravascular coagulation( DIC) (11. 1% ). Laboratory data mainly manifested with cytopenia ( 100% ) , liver dysfunction ( 77. 8% ) , hypofibrinogenemia (62. 8% ), hypertriglyceridemia (71. 1% ) , serum ferritin >500 p,g/L(87. 5% ) , low NK-cell activity(92. 9% ) and hemophagocytosis in bone marrow (100% ). Based on treating underlying diseases and use of corticosteroids and immunosuppressive agents in combination with intravenous immunoglobulins(IVIG) therapy, 14 patients recovered , 19 patients died in the hospital, and other 12 cases give up treatment be cause of exacerbation of the disease.Conclusion There are various underlying diseases and clinical manifestations for HPS. HPS is always extremely dangerous with high mortality. Treatment of underlying diseases, corticosteroids, immunodepressant and IVIG may improve the prognosis of HPS.

Objective To evaluate the role of microRNA130α on rat bone mesenchymal stromal cells(BMSCs)during chondrogenic differentiation.Methods BMSCs were induced to differentiate into chondroeytes by transforming growth factor-β1(TGF-β1)in vitro,immunofluorescence and immunohistochemistry were performed to evaluate MSCs differentiation.RT-PCR was performed to analyze microRNA130α expression at different time points.Results microRNA130α was down-modulated during chondrogenesis after BMSCs been cultured with TGF-β1 for 7 days (P ＜0.05).Conclusion During the early stage of BMSC chondrogenic differentiation,mciroRNA130a expression was specifically repressed,suggesting its role in differentiation of rat bone mesenchymal stromal cells.

Objective To analyze the clinical characteristics of pneumomediastinum in patients with dermatomyositis and polymyositis for demonstrating its pathogenesis and for predicting its prognosis. Methods The clinical records of 96 patients with PM/DM were reviewed, focusing on for perdicting its pneumomediastinum. Five patients with pneumomediastinum are described in detail. Case reports of pneumomediastinum in PM/DM in English publications are reviewed. Results Five DM cases complicated by pneumomediastinum all had lung infections. Twenty-nine cases (including our five cases) of DM/PM with pneumomediastinum have taken methylprednisolone, four cases alive, and six died. Nine cases have taken CsA,seven cases alive and two died. Conclusion The infections was strongly suspected as being responsible for the pneumomediastinum. Methylprednisolone has poor effect. CsA can be an effective therapeutic agent in PM/DM.

Objective To investigate that p53 expression in fibroblast-like synoviocytes (FLS) and its effects on CD4+ T lymphocytes from active rheumatoid arthritis (RA) patients. Methods Human FLS was transfected with p53siRNA and cocultured with CD4+ T lymphocytes from active RA patients. The expression of osteoprotegerin and IL-6 secretion was detected in the transfected FLS. In addition, the expressions of IFN-γ, IL-17, IL-4 and CD25 as well as mRNA levels of IFN-γ RORγt, IL-17 and Foxp3 in cocuhured CD4+ T lymphocytes were also measured. Results IL-6 secretion decreased in p53-inhibited FLS while osteopro-tegerin expression was not altered, p53-inhibited FLS could significantly increase IL-17 and IFN-γ expres-sions. It also upregulated Foxp3 expression though had no effect on CD4+CD25high T lymphocytes. Conclusion p53 expression in FLS regulates Th1 and Th17 cells of RA patients, and therefore participate in the pathogenesis of RA.