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Objective To investigate the effect of glucocorticoid and bisphosphonate on Hedgehog signaling pathway in human bone mesenchymal stem cells (hBMSCs) and bone tissue.Methods ① Bone biopsy test:forty cases of systemic lupus erythematosus (SLE) patients were divided into 2 groups:20 cases in newly diagnosis group,20 cases in the GCs group (the dosage of glucocorticoids was higher than 1 mg·kg-1·d-1).Patients in the GCs group was randomly divided into 2 groups:10 cases in the control group were without anti-osteoporosis treatment,10 cases in treatment group received alendronate 70 mg once a week.All of the patients had bone marrow puncture after24 weeks,and the value of average optical density of Gli1 was tested with immunohistochemistry.② Cell culture:hBMSCs cultured in normal medium were intervened with rh-SHHN and methylprednisolone (10-3 mol/L,10-5 mol/L) after successfully identified.Quantitative polymerase chain reaction (PCR) was used to detectthe mRNA expression of Gli1.The final calcium nodules was detected by Alizarin red staining.hBMSCs cultured in osteogenesis medium were intervened with bisphosphonate and methylprednisolone (10-3 mol/L) after successfully identified.Quantitative PCR was used to detect the expression of Gli1 mRNA.Alizarin red staining was used to detect the final calcium nodules.Comparisons between the two groups were carried out using t-test while the difference analysis of multi-groups were tested by factorial analysis.Results The average optical density of Gli1 in the GCs group (47±7) was less than the newly diagnosed group (61±12) (t=4.442,P＜0.01),and it was less in the control group (51±6) than in the treatment group (42±6) (t=3.701,P=0.002).When normally cultured,high and moderate concentration of methylprednisolone suppressed the mRNA (0.38±0.04; 0.68±0.24) (F=8.748,P＜0.01) expression of Gli1 which was initially stimulated by rh-SHHN (2.01 ±0.38).And the final calcium nodules in groups which contained methylprednisolone were much less than rh-SHHN only group.When hBMSCs were cultured in osteogenesis medium,compared with the methylprednisolone group (0.024±0.011),the expression of Gli1mRNA(0.034-0.006) (t=7.62,P＜0.01) and the final calcium nodules were significantly improved by bisphosphonate.Conclusion High and moderate doses of glucocorticoids inhibit hBMSC osteogenesis by inhibiting Gli1.Low concentration of alendronate can not only stimulate hBMSC osteogenesis differentiation but also can remit the inhibition effects of GCs through the way of Hedgehog.
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Objective To investigate the expression of lipid rafts (LRs) and actin cytoskeleton (F-actin) in the peripheral blood B lymphocytes of patients with systemic lupus erythematosus (SLE).Methods Peripheral blood mononuclear cells (PBMCs) were separated by Ficoll-Hypaque.B lymphocytes were isolated by positive selection from PBMCs.Membrane staining for LRs was achieved with FITC-conjugated cholera toxin B (CTB).The level and distribution of LRs were studied by flow cytometry and confocal microscopy.Staining for F-actin was carried out with Rhodamine phalloidin.The expression of F-actin was analyzed by confocal microscopy.In an in vitro examination,the effect of Leflunomide on lipid rafts in B lymphocytes from SLE was analyzed.Disease carried out was measured using the SLE disease activity index (SLEDAI).Analysis of the enumerical data was performed using ANOVA or paired-samples t test.Correlation was examined by Pearson's rank correlation test.Results The number of CTB-binding lipid rafts in B cell from active SLE patients or from SLE patients in disease remission.who were treated with immunosuppressive drugs was higher than B cells from healthy controls [(59+4)％,(51±5)％,(33±4)％,F=9.21,P=0.001].Confocal microscopy revealed that in B cell from healthy controls,lipid raft was found to be small and uniformly distributed on the plasma membrane.F-actin was found mainly in the cortical region of the cells.This pattern was different from the pattern seen in B cells from patients with SLE,which presented with stronger staining and irregular large clustering of LRs,with a decrease in F-actin levels.In addition,the number of CTB-binding LRs in B cells from the active SLE patients was correlated significantly with the SLEDAI score (r=0.632,P=0.028).Furthermore,thein vitro results showed that leflunomide treatment reduced the number of CTB-binding LRs in B cell from SLE patients [(48±5)％ vs (39±5)％,t=2.29,P=0.048].Conclusion The altered expression of Lipid raft and F-actin can been seen in B lymphocytes in SLE,and modulation of LRs and F-actin expression may be a potential approach in the treatment of SLE.
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Objective To analyze the clinical features of renal involvement associated with Behcet's disease (BD) through 1 case and to make a review of the literature in order to early diagnose and cure in time,thus decrease misdiagnosis and mistreatment.Methods This is a retrospective study.The case was diagnosed with BD and the renal damage was confirmed by renal biopsy.The clinical features and histology features were analyzed.Results The presentation of renal disease was edema,proteinuria and microscopic hematuria.The clinical spectrum of renal BD showed a wide variation.Amyloidosis (AA type),GN (nephritis),and microscopic vascular disease were the main causes of renal BD.Patients with vascular involvement had a high risk of amyloidosis and amyloidosis was the most common cause of renal failure in BD.Conclusion Kidney is one of the organs that can alter the prognosis of the BD,so the screening for renal damage must be done for each patient with this disease.Routine urine analysis and measurement of serum creatinine level are needed for early diagnosis of renal BD.Immunosuppressive drugs can be useful in selected cases.
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Objective To investigate the effect of leflunomide on the superficial costimulatory molecules expression of T lymphocytes in patients with lupus nephritis ( LN ). Methods The peripheral blood mononuclear cells (PBMCs) of female active LN patients and healthy female were separated by density gradient centrifugation, and cultured by phytohemagglutinin (PHA) or leflunomide active metabolites(A771726).The CD28, CD40L, CTLA-4 and LFA-1a expressions on the peripheral blood T lymphocytes were detected by double-colored flow cytometry. The differences of the means were tested by analysis of variance( ANOVA ) and SNK q test. Results Comparing with healthy controls, there were significantly higher expressions of CD28,CD40L, LFA-1a and CTLA-4 on the peripheral blood T cells in active LN patients (CD28:33.4±6.5 vs14.4±3.2; CD40L: 13.2±3.2 vs 5.4±2.3; LFA-1a: 8.5±2.3 vs2.2±1.1; CTLA-4:4.6±1.5 all P＜0.01) as well as CD28 and CD40L expression on the peripheral blood T cells from healthy controls induced by PHA (CD28:26.8±6.7 vs14.4±3.2; CD40L: 13.9±4.9 vs 5.4±2.3 all P＜0.01 ), but not CTLA-4 and LFA-1a expression.However, CD28, CD40L, LFA-1a and CTLA-4 expressions on T cells stimulated by PHA increased in active LN patients(all P＜0.05 ). A771726 could significantly inhibit over-expression of LFA-1a and CD40L on the T cells from active LN patients (CD40L:8.2±2.0 vs13.3±3.2;LFA-1a:5.1±1.3 vs8.5±2.3 all P＜0.01 ), but not CD28 and CTLA-4 expression. A771726 did not inhibit CD28, CD40L, LFA-1a and CTLA-4 expression on the T cells in healthy controls. Furthermore, A771726 could markedly inhibit the over-expression of all of the above molecules induced by PHA on the T cells of active LN patients (all P＜0.05). Conclusion One of the major mechanisms for LEF treatment of LN is that LEF can down-regulate CD40L and LFA-1a expression but not CD28 and CTLA-4 expression on the peripheral blood T cells in active LN patients.
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In order to study the role of the bone marrow-derived mesenchymal stem cells(MSCs)transplanted with or without bone marrow(BM)in the treatment of lupus mice and the effect of MSCs in the onset of systemic lupus erythematosus(SLE).Method Twenty 12-week-old female MRL/lpr mice were randomly divided into four groups,including simple bone marrow transplantation group(SG,BM 1×107),united group-1(UG1,BM 1×107+MSCs 1×104),united group-2(UG2,BM 1×107+MSCs 1×106),the positive control group(PG,no transplantation).BALB/c mice were used as the negative control group(NG,no transplantation).MSCs which were amplified from the bone marrow of male BALB/c mice in vitro were transplanted into the female MRL/lpr mice with or without BM.One month later Y chromosome was detected to confirm if the transplantation was successful or not.The change of weight, white blood cells,urine protein,anti-dsDNA antibody,the pathology and immunofluorescence of renal were observed to evaluate the therapeutic efficacy.Results Y chromosome was detected in all transplanted female mice.Compared with PG,urine protein concentration in SG,UG1 and UG2 significantly decreased 30 days after transplantation(P<0.05).40 days after transplantation,the rite of anti-dsDNA antibodies in sG(0.91±0.27)was still higher than NG,which OD value wag 0.47 s0.10(P<0.05),but there was no statistical difference among UG1(0.76±0.28),uG2(0.73±0.10)and NG(P>0.05).However,50 days after transplantation,there was no marked difference of the tite of anti-dsDNA antibodies in SG(0.55±0.15),UG1(0.57±0.14)and UC2(0.58±0.05)compared with NG(P>0.05).After transplantation there was no vasculitis.no inflammatory cell infiltration in matrix and no obvious intercapillary cells proliferation in the kidney.The immunofluoroscence became negative or weakly positive.Conclusion MSCs transplantation with or without BM Can both improve the pathogenetic condition of MRL/lpr mice.MSCs can accelerate the clearance of anti-dsDNA antibody and promote the restoration of injured organs.We presume that MSCs are important immunological regulation cells in SLE.
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Objective To investigate the effect of thymosin α1 on immun ologic function and cytokines in the abdominal sepsis patients.and naddiscuss the therapeutic function of thymoin α1 in sepsis.Methods Thirty-eight sepsis patients admitted between May 2006 and May 2007 in the Hospital of Weifang Medical University.Patients were divided into the treatment group(20 cases)and control group(18 cases)randomly.Thymosin α1 was used in the treatment group on the basis of infection controlled.The concentration of cytokines(TNF-α,IL-6,IL-10)and the percent age of CD3,CD4,NK cell were observed.Results The concentrations of TNF-α and IL-6 were increased significantly in bothgoups.But it was more prominently in the control group.The concentration of IL-10 and the percent age of CD3,CD4,NK cell increased more significantly in thymosinod treatment group than those in the control group.Conclusions Thymosincd canmodulate the release of TNF-α,IL-6,IL-10 and mitigate the inflammatory reaction,which may improve the immunologic function.