Objective To evaluate the inhibiting effect of selective and non-selective cyclo-oxygenase-2(COX-2) inhibitor on growth of colon cancer cell lines in vitro and its signal transduction pathway. Methods The proliferation of colon cancer cells was determined by MTT assay. COX-2, nuclear factor(NF)-?Bp65, extracellular-signal regulated kinase(ERK), phospho ERK(pERK) protein expressed in colon cancer cell lines were analyzed by Western blot. Activator protein(AP)-1 and NF-?B binding activity influenced by non-steroidal anti-inflammatory was detected by electrophoretic mobility sift assay. Results Aspirin and celecoxib could inhibit the proliferation of HT-29, SW480 and LS174-T cells and showed a dose-dependent manner. Western blot analysis showed that expression of COX-2, pERK, NF-?Bp65 protein in colon cancer cells were down-regulated by celecoxib or aspirin in some degree but not effect in total ERK expression. AP-1 and NF-?B binding activity could be stimulated by 20% fetal calf serum or tumor necrosis factor-?. Both aspirin and celecoxib could inhibit fetal calf serum-induced AP-1 activation in HT-29 and SW480 cells. Celecoxib could also inhibit tumor necrosis factor induced NF-?B binding activity, but aspirin had little effects on SW480 cells. Conclusion NSAIDs are able to potentially inhibit the growth of colon cell lines in vitro and the mechanism may relate to AP-1 and NF-?B signal transduction pathway.

Objective To investigate the adjustment effect of microRNA-155 on CD4+CD25+ regulative T cell (Treg) in peripheral blood of sepsis patients,and to elucidate the role of miR-155 in the pathogenesis of sepsis.Methods A retrospective study was corducted.60 sepsis patients (mild n =20,moderate n=20,and severe n =20)from emergency room or intensive care unit (ICU) of the Fourth Hospital of Jiangsu University were enrolled.20 healthy volunteers were enrolled as controls.Real-time fluorescent quantitation polymerase chain reaction (qRT-PCR) was used to detect the levels of miR-155 and Foxp3 mRNA expressions in peripheral blood.CD4+CD25+ Treg cells in peripheral blood were identified by flow cytometry.Peripheral interleukin-10 (IL-10) was measured by enzyme-linked immunosorbent assay (ELISA).Results The expressions of miR-155,Treg,Foxp3 mRNA and the level of IL-10were higher in the patients with sepsis than those in healthy control group [Treg:(2.89 ± 1.13)％ vs.(2.32 ± 0.91)％,t=10.540,P=0.002; miR-155:1.19 ±0.48 vs.0.80 ±0.33,t=8.605,P=0.006; Foxp3 mRNA:0.18 ±0.08 vs.0.13 ±0.03,t=6.862,P=0.008; IL-10 (ng/L):56.89 ± 17.28 vs.33.24 ± 11.93,t=12.742,P=0.001].These variables were elevated gradually with the elevation of acute physiology and chronic health evaluation Ⅱ (APACHE Ⅱ)score.The expressions of Treg,miR-155,Foxp3 mRNA and the level of IL-10 were (3.05 ± 1.21)％,1.36 ± 0.79,0.21 ±0.10,(62.82 ±21.38) ng/L in severe sepsis group; they were (2.86 ±0.88)％,1.25 ±0.56,0.17 ±0.08,(56.38 ± 19.65) ng/L in moderate group; they were (2.61 ± 0.87)％,0.94 ± 0.52,0.15 ± 0.05,(45.43 ± 14.40) ng/L in mild group.The values showed significant statistical difference among the mild,moderate and severe sepsis groups (all P＜0.01).Above values were significandy higher in non-survival group than those in survival group [Treg:(3.46 ±1.53)％ vs.(2.85 ± 1.03)％,t=14.250,P=0.005; miR-155:1.41 ±0.85 vs.1.16 ±0.76,t=11.875,P=0.006;Foxp3 mRNA:0.24 ± 0.11 vs.0.17 ± 0.09,t=8.795,P=0.001 ; IL-10 (ng/L):65.47 ± 23.58 vs.51.70 ± 16.86,t=16.313,P=0.001].The expression of miR-155 was positively correlated with the expression of CD4+CD25+ Treg and Foxp3 mRNA (r1=0.635,P1=0.007; r2=0.671,P2=0.005).Conclusion The result of this study suggest that miR-155 is involved in the cell proliferation regulation of CD4 +CD25 + Treg cells,and play some role in the immunological dissonance in sepsis.

BACKGROUND:Stromal cel derived factor-1 is a smal molecular protein with a wide range of biological activity that can cause immune cel chemotaxis, and it also has a chemotactic effect on bone marrow stem cels and periodontal ligament cels.
OBJECTIVE:To investigate the effect of stromal cel derived factor-1 with different concentrations on the proliferation of bone marrow stem cels and to probe the best concentration.
METHODS:Bone marrow stem cels from beagle dogs were culturedin vitro and stimulated by different concentrations of stromal cel derived factor-1 (100, 200, 300 μg/L). MTT was used to detect the influence of stromal cel derived factor-1 on the proliferation of bone marrow stem cels so as to screen the best concentration of stromal cel derived factor-1. Then, stromal cel derived factor-1 at the best concentrations was used to intervene the bone marrow stem cels, and MTT was used again to detect the proliferation of bone marrow stem cels.
RESULTS AND CONCLUSION:Stromal cel derived factor-1 at concentrations of 100, 200, 300 μg/L could promote the proliferation of bone marrow stem cels, and the effect was more notable at 200 and 300 μg/Lbut withno significant difference. Therefore, 200 μg/L was considered to be the best concentration of stromal cel derived factor-1 for intervention of bone marrow stem cels. Compared with the blank control group, 200 μg/L stromal cel derived factor-1 could significantly promote the proliferation of bone marrow stem cels. Taken together, stromal cel derived factor-1 can promote the proliferation of bone marrow stem cels, and its best concentration is 200 μg/L.

Objective:To explore the expression of IL-22 in the patients with rheumatoid arthritis,and to define the clinical sig-nificance of IL-22 cells for RA. Methods: A total of 50 RA patients, 15 OA patients and 15 healthy controls were enrolled. The proportion of Th22 cells in peripheral blood and synovial fluid( SF) of RA patients was detected by flow cytometry;the levels of IL-22 in serum and synovial fluid of RA patients were detected by ELISA. The clinical parameters of disease activity were assessed including ESR,RF,CRP,DAS28,anti CCP and the degree of bone erosions determined by X-rays. Pearson correlation analysis,t test and Kruskal-Wallis H test were used for statistical analysis. Results:The proportion of Th22 cells in RA patients was higher than that of OA patients (t=2. 290 ,P=0. 021) and healthy controls(t=2. 524,P=0. 015). IL-22 levels in RA patients were higher than that of OA patients (t=2. 560,P=0. 014) and healthy controls(t=2. 768,P=0. 009). IL-22 in the RF positive group were higher than that of RF negative group(t=2. 322,P=0. 035). IL-22 in the anti-CCP antibody positive group were higher than that of anti-CCP antibody negative group (t=2. 504,P=0. 015). The levels of IL-22 were correlated positively with ESR,RF,DAS28(r=0. 312,0. 314,0. 332,P<0. 05). The levels of IL-22 were increased gradually along with the aggravation of bone erosions. There were statistical significant difference among different X-ray stagings(H=9. 14,χ20.05(3)=7. 81,H> χ20.05(3),P<0. 05). Serum IL-22 levels in the RA patients with joint effusion were higher than that of without(t=2. 587,P=0. 012). The levels of IL-22 in SF were higher than that in serum(t=2. 668,P=0. 011),and had no correlation with the proportion of Th22 cells in SF. Conclusion: The expression of IL-22 in serum and joint effusion of RA patients increase. The level of IL-22 may be useful marker for assessment of disease activity and finding of bone erosions. Therapeutic targeting of IL-22 may be valuable for the intervention of RA.

OBJECTIVETo purify the arsenic-binding proteins (As-BP) in hamster plasma after a single oral administration of arsenite (iAs(III)).

METHODSArsenite was given to hamsters in a single dose. Three types of HPLC columns, size exclusion, gel filtration and anion exchange columns, combined with an inductively coupled argon plasma mass spectrometer (ICP MS) were used to purify the As-BP in hamster plasma. SDS-PAGE was used to confirm the arsenic-binding proteins at each purification step.

RESULTSThe three-step purification process successfully separated As-BP from other proteins (ie, arsenic unbound proteins) in hamster plasma. The molecular mass of purified As-BP in plasma was approximately 40-50 kD on SDS-PAGE.

CONCLUSIONThe three-step purification method is a simple and fast approach to purify the As-BP in plasma samples.

Administration, Oral ; Animals ; Arsenic ; blood ; Arsenites ; administration & dosage ; pharmacokinetics ; Carrier Proteins ; blood ; Chromatography, High Pressure Liquid ; methods ; Cricetinae

In our previous work, we found that trivalent dimethylarsinous acid (DMA(III)) have high affinity binding to cysteine residue 13 of rat hemoglobin. However, it is still unknown why arsenic intermediate metabolite DMA(III) has high binding affinity for Cysl3 but not for other cysteine residues 93, 140, 111 and 125. In order to better understand the molecular mechanism of DMA(III) with rat hemoglobin, we have done current study. So, SD rats were divided into control and arsenic-treated groups randomly. Arsenic species in lysate of red blood cells were analyzed by HPLC-ICP-MS, and then determined by a hybrid quadrupole TOF MS. In addition, trivalent DMA(III) binds to different cysteine residues in rat hemoglobin alpha and beta chains were also simulated by Molecular Docking. Only Cys13 in alpha chain is able to bind to DMA(III) from the experiment results. Cys13 of alpha chain in rat hemoglobin is a specific binding site for DMA(III), and we found that amino acids compose pockets structure and surround Cys13 (but not other cysteine residues), make DMA(III) much easy to bind cysteine 13. Taken together, the DMA(III) specific binding to Cys13 is related to spatial structure of Cys13.

ObjectiveTo unify the definitions of colonoscopic characteristics of Crohn disease (CD) and intestinal tuberculosis ( ITB),and to evaluate colonoscopic and clinical features in the differential diagnosis of CD and ITB.MethodsA collaborative group composed of 10 experts from 5 hospitals voted to identify and confirm the colonoscopic characteristics.Clinical and colonoscopic characteristics were analyzed,thereafter,characteristics were scored based on different diagnostic specificity.ROC curve was used for determining the cutoff point to differentiate CD from ITB.ResultsFirstly,standard endoscopic images and descriptions were determined.Secondly,colonoscopic parameters which were significantly different between the CD and ITB patients included the follows:involvement of more than four intestinal segments,anorectal involvement,longitudinal ulcers,cobblestone appearance and transverse ulcers.Clinical findings which were significantly different between the CD and ITB patients included active pulmonary tuberculosis,PPD-test strong positive,anal fistula/perianal abscess and extra-intestinal manifestations in CD.4.4％(6/136) patients were confirmed by histological evidence of caseating granulomas.By using our scoring system,39.7％ (54/136) confirmed diagnoses and 18.4％ (25/136) suspected diagnoses were made in patients without histological evidence.ConclusionIdentification of colonoscopic characteristics and unification of the colonscopic diagnostic criteria were helpful in the differential diagnosis between CD and ITB.The differential diagnosis rate could he improved by using the scoring system.Half cases could not be confirmed even with combined pathology and the scoring system,so a more comprhensive scoring system would be warranted.

Objective To investigate the effect of macrophage FRβ expression on bleomycin induced pulmonary fibrosis(PF) in mice.Methods The rats were divided into the normal group,control group and experimental group,6 cases in each group.The mice were performed the PF induction.The experimental group was treated with immunotoxin,the control group was given the contrast protein and the normal group was not treated.The mouse left lung was used for histological analysis,and the right lung was used for hydroxyproline analysi s.The effect of macrophage FRβ expression on bleomycin induced pulmonary fibrosis in mice and the pulmonary macrophage FRβ expression in the patients with idiopathic pulmonary fibrosis(UIP) were detected.Results The macrophage FRβ expression mainly existed in the patients with UIP and pulmonary fibrosis area of PF mice induced by bleomycin;the survival rate was significantly increased by giving the mice immunotoxin with nose(P=0.003),and the level of total hydroxyproline and fibrosis of PF mice induced by bleomycin was decreased(P=0.009,0.014);immunohistochemistry results showed that immunotoxin could reduce the cells number of lung tumor necrosis factor(TNF)-α,chemotactic CCL2 and CCL12 cells inPF mice induced by bleomycin(P=0.000).Conclusion The FRβ expression of macrophages plays a pathogenic role in IPF,and the targeted therapy of FRβ expression in macrophages may be an effective method for the treatment of IPF.

This study was conducted to evaluate the growth and NAG-1 gene expression effected by Non-steroidal anti-inflammatory drug (NSAID) on colon cancer cell lines in vitro. The proliferation of colon cancer cells were determined by MTT assay and COX-2 protein expression were detected by Western blot. Total RNA was isolated from three kinds of colon cancer cell lines; the expressions of NAG-1 mRNA in the cells treated with or without NSAIDs were assessed by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) assay. Celecoxib, meloxicam and aspirin were able to inhibit the growth of HT-29, SW480 and LS174-T cells in dose-dependent manner. COX-2 protein expressed in HT-29 and LS174-T, but not in SW480 cells. All of colon cancer cells expressed NAG-1 gene and the level of LS174-T was lower than that of the other two cell lines. NAG-1 expression was increased by treatment with some NSAIDs in all three kinds of colon cancer cells. NSAIDs were able to potentially inhibit the growth of colon cell lines. Induction of NAG-1 gene expression by NSAID was not consistent with COX-2 expression.
Anti-Inflammatory Agents, Non-Steroidal ; pharmacology ; Antineoplastic Agents ; pharmacology ; Aspirin ; pharmacology ; Celecoxib ; Cell Proliferation ; drug effects ; Cyclooxygenase 2 ; metabolism ; Gene Expression Regulation, Neoplastic ; HT29 Cells ; Humans ; Neoplasm Proteins ; metabolism ; Pyrazoles ; pharmacology ; RNA, Messenger ; metabolism ; Sulfonamides ; pharmacology ; Thiazines ; pharmacology ; Thiazoles ; pharmacology

Objective To investigate the clinical and pathological characteristics of Henoch-Sch?nlein purpura nephritis with diffuse capillary endothelial cell proliferation as pathological manifestation. Methods The clinical manifestations and pathology of capillary endothelial proliferative purpura nephritis (DEP-HSPN) diagnosed by renal biopsy were retrospectively analyzed in 50 children in recent 5 years. Results The pathological lesions in 50 cases included simple DEP-HSPN in 11 cases (7 males and 4 females) and capillary endothelial cell proliferation combined with crescents formation (non-simple DEP-HSPN) in 39 cases (27 males and 12 females). There was no significant difference in the course of disease and age between the two groups (P>0.05). The clinical type of 11 cases of simple DEP-HSPN was type Ⅲ. In 39 cases of non-simple DEP-HSPN, 16 cases were type Ⅲ and 23 cases were type Ⅴ. All of the children had hematuria and proteinuria. The incidence of gross hematuria, urine red blood cell count, 24 h urine protein, and serum creatinine levels in children with non-simple DEP-HSPN were significantly higher than those in simple DEP-HSPN group, but the plasma albumin level was significantly lower than that in simple DEP-HSPN group. It was easy to have crescent formation in DEP-HSPN, and the rate of crescent formation was 11.1％ (5.0％-27.6％). The incidence of segmental lesions and renal tubular interstitial damage was low. All children had non simple IgA deposits in the mesangial area. In the 50 children treated for 1 year, 22 had complete remission, 28 had asymptomatic hematuria, and none had active nephropathy and renal insufficiency. In 32 cases of non-simple DEP-HSPN, the 24 h urinary protein, plasma albumin level, and the incidences of gross hematuria and microscopic hematuria were statistically different before treatment and 1, 3, 6, 12 months after treatment (P<0.01). The 24 h urine protein and gross hematuria gradually decreased with the prolongation of treatment, while the level of plasma albumin was gradually increased. Conclusions DEP-HSPN is characterized by gross hematuria and proteinuria. The onset is acute and it is easy to have crescent formation. When combined with crescent formation, the clinical symptoms are more severe. The combination of strong immunosuppressive agents and long-term sequential follow-up treatment is effective in acute stage. The prognosis is good.