Thalidomide and tuberculosis.

The anti-inflammatory and immunomodulatory effects of thalidomide have led scientists to explore its clinical therapeutic values. Thalidomide is now being considered as an adjuvant treatment for tuberculosis. This literature review examines the drug's mechanism of action and clinical applications. Thalidomide affects cytokine production and T lymphocyte proliferation. It appears that thalidomide suppresses TNF-alpha production by macrophages and thereby reduces inflammatory response. Thalidomide elevates the IFN-gamma level and modulates several other cytokines as well, noteworthily IL-2 and IL-12. Thalidomide costimulates T lymphocytes, with greater effect on CD8+ than on CD4+ T cells. This finding is important, since CD8+ T cells have been shown to be contributory to the protective immune response to Mycobacterium tuberculosis infection. The clinical application of thalidomide as part of standard tuberculosis therapy is inconclusive amid variability among reports. However, thalidomide has been shown to be an effective adjuvant for tuberculosis patients complicated with severe inflammatory reaction or wasting conditions.     

The effects of thalidomide on chemotactic migration of multiple myeloma cell lines

We examined the effect of thalidomide and dexamethasone on the migration of multiple myeloma (MM) cell lines, U266, RPMI8226, and NCI-H929, using chemotaxis chamber plates. U266 underwent chemotactic migration in response to stromal-cell derived factor-1 alpha (SDF-1alpha), and other cell lines underwent random migration in response to SDF-1alpha or monocyte chemotactic protein-1 alpha. Following preincubation with 1 mug/ml thalidomide, the cell lines showed reduced migratory capacity in response to SDF-1alpha. Concerning the corresponding receptors, CXC chemokine receptor 4 was detected only on the surface of U266, by flow cytometry, whereas chemokine (C-C motif) receptor 2 was not detected on all three cell lines. Moreover, decreased migration by thalidomide was not accompanied by altered expression of the corresponding receptors of each cell line. This is the first report to show the effects of thalidomide on the migration of MM cell lines. The results suggest that the inhibition of chemotactic migration might be one of the mechanisms of the success of thalidomide in controlling MM.     

Preclinical and clinical assessment of the safety and potential efficacy of thalidomide in heart failure

BACKGROUND: Inflammatory mediators, especially tumor necrosis factor (TNF), have been implicated in heart failure (HF). Thalidomide has anti-inflammatory properties and selectively inhibits TNF. Thus far, thalidomide or thalidomide analogues have not been evaluated in patients with heart failure. METHODS: Thalidomide was assessed in preclinical and clinical studies. First, isolated cardiac myocytes were pretreated with thalidomide or thalidomide analogues, and TNF production was assessed after lipopolysaccharide (LPS) provocation. Second, to determine the safety and potential efficacy of thalidomide, an open-label dose escalation safety study was conducted in seven patients with advanced heart failure. RESULTS: Thalidomide and thalidomide analogues inhibited LPS-induced TNF biosynthesis in cardiac myocytes in a dose-dependent manner. Thalidomide analogues had a greater inhibitory effect on TNF production than did thalidomide. In patients with advanced HF, thalidomide was safe and potentially effective when used at lower doses. However, dose-limiting toxicity was observed in two patients. There was a significant increase in the 6-minute walk distance and a trend toward improvement in left ventricular ejection fraction and quality of life after 12 weeks of maintenance therapy with thalidomide. CONCLUSIONS: Taken together these results suggest that thalidomide or its derivatives may be useful in selected patients with HF. This potential needs to be studied in larger clinical trials.     

高迁移率族蛋白-1在类风湿关节炎患者单核细胞中的移位和释放及沙立度胺对其的影响

目的 探讨类风湿关节炎(RA)患者外周血单核细胞高迁移率族蛋白-1(HMGBl)的释放和细胞内定位及沙立度胺对其的影响.方法 抽取19例RA患者和20例健康对照者外周静脉血5 ml,Ficoll密度梯度离心法分离外周血单核细胞,分别用100 ng/ml TNFα、100 ng/ml TNFα+40 μ/md沙立度胺处理细胞后,Western blot检测外周血单核细胞HMGB1的释放;细胞免疫荧光法观察外周血单核细胞中HMGB1的胞内定位.结果 (1)在无刺激因素作用下,RA患者外周血单核细胞培养液的上清中HMGBl蛋白水平较健康对照组显著增高(P<0.05).100 ng/ml TNα并不增加RA患者外周血单核细胞中HMGBl的释放,40μg/ml沙立度胺能明显降低RA患者外周血单核细胞HMGBl的释放(P<0.05).(2)RA患者外周血单核细胞中HMGB1主要分布于胞核.100 ng/m1TNFα刺激外周血单核细胞24 h后HMGB1向胞质移位,40 μg/m1沙立度胺能明显抑制TNFα所致的HMGBl胞质移位.结论 RA患者外周血单核细胞在TNFα刺激下HMGB1从核内移位至胞质,最后释放到细胞外,沙立度胺能抑制HMGB1的胞质移位和释放.     

Spontaneous production of mediators by mononuclear human blood cells (author's transl)

Human mononuclear blood cells incubated in vitro without any intended stimulus produce lymphokine-like activities. Separated lymphocytes release a leukocyte migration stimulatory factor spontaneously. The spontaneous production of a leukocyte migration inhibitory factor in unstimulated cultures during the first 20 hours needs for a cooperation between lymphocytes and monocytes. As well lymphocytes as enriched monocytes elaborate leukocyte chemotactic activity. --The spontaneous activation is caused by the increasing acidity of the culture medium following metabolic processes. The dextran sedimentation and the Ficoll content of the separation solution were without importance for the stimulation.     

沙利度胺对类风湿关节炎T淋巴细胞的免疫调节作用

目的 探讨沙利度胺(Thd)对类风湿关节炎(RA)患者外周血T淋巴细胞的免疫调节作用.方法 在植物血凝素(PHA)刺激下,RA患者外周血T淋巴细胞与不同浓度的Thd(2.5、25、100、300、500μg/ml)共同培养.用四甲基噻唑蓝(MTT)比色法检测T淋巴细胞的增殖,用流式细胞术检测T淋巴细胞的凋亡和T淋巴细胞CD152、CD28的表达情况,用Real-time聚合酶链反应(PCR)检测T淋巴细胞的自细胞介素(IL)-6、IL-10、肿瘤坏死因子(TNF)-α mRNA表达水平.结果 在体外,与阴性对照组相比,500μg/ml,Thd组显著抑制T淋巴细胞的增殖,促进T淋巴细胞早期凋亡,抑制T淋巴细胞CD3+CD28+表达,促进CD8+CD152+表达,而≤300μg/ml,对T淋巴细胞未见明显影响.与对照组相比,(100～500)μg/ml剂量组可抑制T淋巴细胞IL-6、TNF-α mRNA表达,(2.5～500)μg/ml可促进IL-10 mRNA的表达.结论 Thd可能通过影响RA患者外周血T淋巴细胞增殖、早期凋亡、CD3*CD28+和CD8+CD152+表达,以及IL-6、IL-10、TNF-α mRNA表达,反转RA患者的免疫平衡失控.     

The future role of thalidomide in multiple myeloma

Treatment for multiple myeloma typically is chemotherapy, high-dose therapy with stem cell transplantation or radiation. Thalidomide shows promise, but to improve knowledge and understanding of this drug, more clinical data is required. Workshops designed to propose new studies providing much needed data on the use of thalidomide as first-line therapy, as maintenance therapy and as treatment for refractory or relapsed disease are reported here. Thalidomide has been shown to be effective in previously untreated patients as monotherapy (36% remission), although greater remission is seen in combination therapy, for example, thalidomide with dexamethasone (72% remission). Thalidomide is an interesting maintenance therapy. It has the advantages of being effective in the bone marrow and having a long-lasting effect but suffers because of its significant side-effect profile; however, it is tolerated by most patients. The adverse effects are thought to be dose-limiting, and most are seen in all patients at the beginning of treatment. Further clinical evidence is needed to determine the effectiveness of thalidomide as maintenance therapy in patients with multiple myeloma, having been shown to be active against refractory multiple myeloma, even in patients who relapse after high-dose chemotherapy. The workshop acknowledged the problems associated with designing a phase III study, and it is clear that many areas need to be addressed in the use of thalidomide for the treatment of multiple myeloma.     

Thalidomide suppresses Up-regulation of human immunodeficiency virus coreceptors CXCR4 and CCR5 on CD4+ T cells in humans

Concurrent infection in patients with human immunodeficiency virus (HIV) infection increases the expression of HIV coreceptors CXCR4 and CCR5. Thalidomide has beneficial effects in a number of HIV-associated diseases. The effect of thalidomide on CXCR4 and CCR5 expression on CD4+ T cells was determined. Thalidomide produced a dose-dependent inhibition of lipopolysaccharide (LPS)-induced up-regulation of CXCR4 and CCR5 in vitro. Antibody to tumor necrosis factor-alpha (TNF-alpha) also attenuated the LPS-induced HIV coreceptor up-regulation, which was not further reduced by thalidomide. Thalidomide (400 mg) was orally administered to 6 men, and their blood was stimulated ex vivo with LPS, staphylococcal or mycobacterial antigens, or antibody to CD3 or CD28 cells. All stimuli induced up-regulation of HIV coreceptors, which was reduced after ingestion of thalidomide. Thalidomide may be beneficial in the treatment of intercurrent infections during HIV infection by reducing the up-regulation of CXCR4 and CCR5 expression on CD4+ T cells induced by bacterial and mycobacterial antigens, by a mechanism that involves inhibition of TNF-alpha.     

Thalidomide in treatment of connective diseases and vasculities

Thalidomide is an immunomodulatory, anti-inflammatory and anti-angiogenic drug. Thalidomide exerts its effects by decreasing circulating CD4 positive T-cells and stimulating CD8 positive T-cells, by increasing the number of Natural Killer cells and T-helper 2 cells. Thalidomide also inhibits proliferation of stimulated T-cells and leukocyte chemotaxis. It modifies a number of integrin receptors and other leukocytic surface receptors and down-modulates cell-adhesion molecules involved in leukocyte migration. It has been demonstrated that thalidomide inhibits TNFalpha, IL-5, IL-6, IL-8, IL-12 production and increases production of IL-2, IL-10 and INFgamma. Moreover thalidomide plays an important role in inhibition of VEGF and FGF-2 mediated angiogenesis. Although the exact mechanism of action is not fully understood and only limited treatment opinions exist, thalidomide plays a role also in connective diseases and vasculities. Thalidomide has been seen efficacious in the treatment of cutaneous disorders in patients with systemic lupus erythematosus and in mucocutaneous disease in Behcet's disease with a not dose-dependent response, even if it should be restricted to selected patients because of its important side effects.     

Patterns of leukocyte chemotaxis to bile after liver transplantation

The role of chemotactic factors in the recruitment of leukocytes to human liver allografts was assessed by studying the effect of bile from transplant recipients on the chemotaxis of cells from normal subjects. Bile samples taken 2-3 days before clinical rejection were more chemotactic for lymphocytes than samples taken during rejection (p less than 0.01), during stable graft function (p less than 0.001), and from nontransplant patients (p less than 0.007). During clinical rejection there was an increase in bile chemotactic activity for both monocytes and neutrophils compared with samples from stable patients (monocytes: p less than 0.001; neutrophils: p less than 0.001) and nontransplant patients (monocytes: p less than 0.001; neutrophils: p less than 0.001). In serial studies chemotactic activity for lymphocytes reached a peak 1-3 days before the onset of clinical rejection, whereas maximum chemotactic activity for monocytes and neutrophils occurred at the time of rejection, when lymphocyte chemotaxis was decreasing. These results suggest that chemotaxis may be important in the recruitment of inflammatory cells to liver allografts and that chemotactic factors for lymphocytes, which appear in bile before clinical rejection, may be critical in the pathogenesis of rejection.     

Thalidomide reduces tumour necrosis factor alpha and interleukin 12 production in patients with chronic active Crohn's disease.

BACKGROUND: Thalidomide improves clinical symptoms in patients with therapy refractory Crohn's disease, as shown in two recent studies. The mechanism of this effect however is still unknown. Suppression of tumour necrosis factor alpha (TNF-alpha) by thalidomide has been suggested as a possible mechanism. However, effects on other cytokines have not been adequately investigated. AIM: The aim of our study was to investigate the effects of thalidomide on cytokine production in patients with inflammatory bowel disease (IBD). METHODS: Ten patients with therapy refractory IBD (nine Crohn's disease, one ulcerative colitis) received thalidomide 300 mg daily in a 12 week open label study. Production of TNF-alpha, interleukin (IL)-1 beta, IL-6, and IL-12 was investigated in short term cultures of stimulated colonic lamina propria mononuclear cells (LPMC) and peripheral blood monocytes (PBMC) before and after 12 weeks of treatment. LPMC were also cultured with graded doses of thalidomide. RESULTS: Three patients discontinued treatment because of sedative side effects. In the other patients, disease activity decreased significantly, with four patients achieving remission. Production of TNF-alpha and IL-12 decreased during treatment with thalidomide: LPMC (TNF-alpha: 42.3 (8.3) pg/ml v 16.4 (6.3); IL-12: 9.7 (3.3) v 5.0 (2.5); p<0.04) and PBMC (TNF-alpha: 62.8 (14.6) v 22.5 (9.2); p<0.02). Production of IL-1 beta and IL-6 did not change significantly. Culturing of LPMC with thalidomide showed a dose dependent decrease in TNF-alpha and IL-12 production. CONCLUSION: The clinical effects of thalidomide in Crohn's disease may be mediated by reduction of both TNF-alpha and IL-12.     

Role of vascular endothelial growth factor in angiodysplasia: an interventional study with thalidomide

Background and Aim: The pathogenesis of angiodysplasia is still not fully understood and effective therapy is not available. Thalidomide was reported to be effective in the treatment of angiodysplasia, but the mechanisms underlying its activity are, as yet, unknown. We aimed to investigate the expression of vascular endothelial growth factor (VEGF) in angiodysplasia tissues, and the role of hypoxia‐inducible factor‐1α (HIF‐1α) and basic fibroblast growth factor (bFGF) on VEGF expression in human umbilical vein endothelial cells (HUVEC). Additionally, we aimed to study the role of thalidomide in these parameters. Methods: Immunohistochemistry was performed to visualize VEGF in angiodysplasia lesions. HUVEC were incubated under hypoxic conditions or in the presence of bFGF. Effects of exposure to thalidomide were studied. Cell growth was assessed in methylthiazolyte‐trazolium assays. Enzyme‐linked immunosorbent assays and real‐time polymerase chain reaction were performed to assess the expression of VEGF at protein and mRNA levels. Western blot was performed to detect the expression of HIF‐1α under hypoxic conditions. Results: VEGF was strongly expressed in 75% of patients with angiodysplasia lesions, as compared to expression in patients without angiodysplasia lesions. VEGF was also induced in HUVEC under hypoxic conditions (P < 0.05). bFGF was found to stimulate the proliferation of HUVEC and enhance the expression of VEGF. Thalidomide suppressed bFGF‐induced proliferation significantly and decreased VEGF expression, both at the protein and mRNA levels. Thalidomide also inhibited HIF‐1α in a dose‐dependent manner (P < 0.05). Conclusions: VEGF may play an important role in the pathogenesis of angiodysplasia. Thalidomide can suppress VEGF, either induced by HIF‐1α or bFGF.     

Chemotactic activity of recombinant human granulocyte colony- stimulating factor

Recombinant human granulocyte colony-stimulating factor (rhG-CSF) induced migration across polycarbonate filters of human polymorphonuclear leukocytes (PMN). rhG-CSF was active in inducing PMN migration at concentrations greater than or equal to 10 to 100 U/mL (7 to 70 ng/mL). rhG-CSF did not contain appreciable levels of endotoxin contamination as assessed by Limulus amebocyte assay, and Polymixin B did not affect the chemotactic activity of rhG-CSF. A monoclonal anti-G- CSF antibody blocked the induction of migration by G-CSF, thus establishing that the cytokine was responsible for the activity of the recombinant preparation. Checkerboard analysis was performed by seeding different concentrations of G-CSF above and/or below the filter and revealed that the migratory response to this cytokine was best observed in the presence of a positive concentration gradient between the lower and upper compartments of the chamber, thus indicating an actual chemotactic effect. When different migrating cells were examined, rhG- CSF was inactive on large granular lymphocytes and endothelial cells under conditions in which appropriate reference attractants were active. In contrast, rhG-CSF elicited a chemotactic response in monocytes inhibited by specific antibody. Thus, G-CSF is a chemotactic signal for phagocytes. This cytokine, when produced at inflammatory sites, may contribute to the recruitment of phagocytes from the blood compartment to amplify resistance against certain noxious agents.     

Autoantigens signal through chemokine receptors: uveitis antigens induce CXCR3- and CXCR5-expressing lymphocytes and immature dendritic cells to migrate

We tested the hypothesis that interaction between autoantigens and chemoattractant receptors may be an important step in the development of autoimmunity. The retinal autoantigens S-antigen (S-Ag) and interphotoreceptor retinoid binding protein (IRBP) can induce autoimmune uveitis in rodent models. We evaluated the chemotactic activity of S-Ag and IRBP and found that both induced migration of human and mouse immature dendritic cells (iDCs) and lymphocytes, but not neutrophils, monocytes, or mature DCs. Cross-desensitization studies and single-receptor transfected cells revealed that subfamily of alpha chemokine receptors CXCR5 and CXCR3 mediated the chemotactic effect of IRBP, while only CXCR3 was required for the chemotactic response to S-Ag. Examination of the relationships between chemoattraction and the ability to elicit pathology at the protein or peptide levels in the mouse uveitis model revealed dissociation of the capacity to induce uveitis, lymphocyte proliferation, and chemoattraction. These studies suggest that IRBP and S-Ag can initiate innate and, in sensitive individuals, adaptive immune response by attracting iDCs and T and B cells expressing CXCR3 and CXCR5.     

Thalidomide induces mucosal healing in Crohn＇s disease： Case report

Crohn’s disease is a chronic inflammatory disorder of the gastrointestinal tract that is defi ned by relapsing and remitting episodes. Tumor necrosis factor alpha (TNF-α) appears to play a central role in the pathophysiology of the disease. Standard therapies for inflammatory bowel disease fail to induce remission in about 30% of patients. Biological therapies have been associated with an increased incidence of infections, especially infection by Mycobacterium tuberculosis (Mtb). Thalidomide is an oral immu...     

Plasminogen-mediated matrix invasion and degradation by macrophages is dependent on surface expression of annexin II

Genetic evidence demonstrates the importance of plasminogen activation in the migration of macrophages to sites of injury and inflammation, their removal of necrotic debris, and their clearance of fibrin. These studies identified the plasminogen binding protein annexin II on the surface of macrophages and determined its role in their ability to degrade and migrate through extracellular matrices. Calcium-dependent binding of annexin II to RAW264.7 macrophages was shown using flow cytometry and Western blot analysis of EGTA eluates. Ligand blots demonstrated that annexin II comigrates with one of several proteins in lysates and membranes derived from RAW264.7 macrophages that bind plasminogen. Preincubation of RAW264.7 macrophages with monoclonal anti-annexin II IgG inhibited (35%) their binding of 125I-Lys-plasminogen. Likewise, plasmin binding to human monocyte-derived macrophages and THP-1 monocytes was inhibited (50% and 35%, respectively) when cells were preincubated with anti-annexin II IgG. Inhibition of plasminogen binding to annexin II on RAW264.7 macrophages significantly impaired their ability to activate plasminogen and degrade [3H]-glucosamine-labeled extracellular matrices. The migration of THP-1 monocytes through a porous membrane, in response to monocyte chemotactic protein-1, was blocked when the membranes were coated with extracellular matrix. The addition of plasminogen to the monocytes restored their ability to migrate through the matrix-coated membrane. Preincubation of THP-1 monocytes with anti-annexin II IgG inhibited (60%) their plasminogen-dependent chemotaxis through the extracellular matrix. These studies identify annexin II as a plasminogen binding site on macrophages and indicate an important role for annexin II in their invasive and degradative phenotype.     

The chemoattraction of lymphocytes by rheumatoid arthritis - synovial fluid is not dependent on the chemokine receptor CCR5

OBJECTIVE: The objective was to study the potential role of the chemokine receptor CCR5 in the chemoattraction of lymphocytes by rheumatoid arthritis synovial fluid (RA-SF). METHODS: The expression of the CCR5 receptor was studied by flow cytometry. Chemotaxis of peripheral blood lymphocytes in response to RA-SF was analyzed on transmigration chambers. Chemotaxis of immortalized lymphocytes from individuals homozygous for the Delta32 deletion of the CCR5 gene (CCR5-/-) was analyzed. The effect of a neutralizing anti-CCR5 antibody on the migration of CCR5+/+ cells was also studied. RESULTS: We confirmed an increase in the proportion of CCR5-expressing lymphocytes in RA-SF and a preferential migration of CCR5+ lymphocytes toward RA-SF in vitro. CCR5-/- lymphocytes showed decreased chemotactic responses to the chemokine MIP-1beta but not to RA-SF. The chemotactic responses of CCR5+/+ lymphocytes to RA-SF were not modified by anti-CCR5 neutralizing antibody. CONCLUSIONS: We confirm a preferential accumulation of CCR5-expressing lymphocytes into RA-SF. However, the chemotactic responses of lymphocytes to RA-SF were not dependent on a functional CCR5 receptor, suggesting that CCR5 is a marker of a lymphocyte subset rather than a specific mediator of chemotactic responses to chemokines in RA-SF.     

Thalidomide--effect on T cell subsets as a possible mechanism of action

Thalidomide is the drug of choice in the erythema nodosum leprosum (ENL) type of lepra reaction. Lately it has been used successfully in other diseases, such as discoid lupus erythematosus, actinic prurigo, Behcet's disease, etc. However, its mechanism of action remains unknown. In patients for whom thalidomide provided relief in their disorder, the proportions of T lymphocytes and their subsets in peripheral blood were assessed by means of monoclonal antibodies. Three lepromatous leprosy patients with ENL had their T helper populations significantly increased after thalidomide therapy. A 14-year-old girl with Behcet's syndrome showed a consistent decrease in Ia+ cells throughout her three-month course of thalidomide therapy. The same findings were observed in two patients with the actinic prurigo type of polymorphous light eruption. From these results, we conclude that thalidomide may act as an immunomodulating agent on T cell subsets.