Differences in autoimmunity factors based on the activity of thromboangiitis obliterans.

OBJECTIVE: The treatment of Buerger's disease (BD) presents a medical problem as its etiology is still unclear. In this study, our objective was to evaluate the serum levels of autoimmune markers in patients with different clinical features of BD. METHODS: In this study, 80 BD patients were categorized in three groups using a cross-sectional design: migratory thrombophlebitis, cold sensitivity, and skin discoloration (mild symptoms); chronic ulcers, claudication, and burning pain of the feet at night (moderate symptoms); pain at rest and spontaneous gangrene (severe symptoms). Enzyme-linked immunosorbent assay was performed to measure antibodies against immunoglobulin M rheumatoid factor (IgM RF), anti-nuclear antibodies (ANA), antibodies against cyclic citrullinated peptide (anti-CCP), antiphospholipid antibodies (APA), anti-cardiolipin antibodies (ACLA), anti-double stranded DNA (anti-dsDNA), and extractable nuclear antigen (ENA) profile. RESULTS: Patients with severe symptoms showed the lowest age (p=0.031), ESR (p<0.001), and highest prevalence of ischemia (p<0.001). In all the patients, the serum levels of ANA and IgM RF were higher than 1 U and 15 IU/mL, respectively. However, the progression of the disease from mild to moderate did not affect these markers significantly (p>0.05). Other markers were negative in patients with BD. CONCLUSION: The findings of this study indicate that BD may closely be correlated to transient autoimmune phenomena, despite the fact that further research is required to investigate how transient unspecific autoimmune reactions contribute to the BD pathogenesis.

Thromboangiitis obliterans plasma-derived exosomal miR-223-5p inhibits cell viability and promotes cell apoptosis of human vascular smooth muscle cells by targeting VCAM1.

Background: Exosomes-encapsulated microRNAs (miRNAs) have been established to be implicated in the pathogenesis of different diseases. Nevertheless, circulating exosomal miRNAs of thromboangiitis obliterans (TAO) remains poorly understood. This study aimed to explore the effects of exosomal miRNAs associated with TAO on human vascular smooth muscle cells (HVSMCs).Methods: The exosomes were isolated from the plasma of TAO patients and normal controls and then were sent for small RNA sequencing. Differentially expressed miRNAs (DE-miRNAs) were identified by bioinformatics analysis and were confirmed by RT-qPCR. After that, PKH67 staining was used to label exosomes and co-cultured with HVSMCs. Cell viability and apoptosis were, respectively, tested by CCK-8 assay and flow cytometry. Finally, dual-luciferase reporter assay was used to confirm the downstream targets of miR-223-5p.Results: A total of 39 DE-miRNAs were identified between TAO patients and normal controls, of which, miR-223-5p was one of the most significantly up-regulated miRNAs. TAO plasma-derived exosomes or miR-223-5p mimics inhibited cell viability of HVSMCs and promoted cell apoptosis. The pro-apoptotic effect of TAO plasma-derived exosomes was alleviated by miR-223-5p inhibitor. Additionally, the expressions of VCAM1 and IGF1R were down-regulated by exosomes and miR-223-5p mimics, and were abrogated by miR-223-5p inhibitor. Dual-luciferase report showed that VCAM1 was the target of miR-223-5p.Conclusions: Our findings imply that circulating exosomal miR-223-5p may play an essential role in the pathogenesis of TAO, and provide a basis for miR-6515-5p/VCAM1 as novel therapeutic targets and pathways for TAO treatment.

ADAM10 inhibits the interaction between IL-17 and HMGB1 in Buerger's disease.

OBJECTIVE: Buerger's disease is a rare disease that causes critical limb ischemia; however, the underlying pathophysiological mechanism remains unclear. Therefore, we investigated the interaction between interleukin (IL)-17 and high-mobility group protein B 1 (HMGB1) and determined whether A disintegrin and metalloproteinase 10 (ADAM10) inhibit this interaction. PATIENTS AND METHODS: The study population included 15 patients with Buerger's disease and 10 healthy donors without a history of giving peripheral blood samples. Cytokine levels were measured using a luminex multiplex assay in plasma. Flow cytometry was used to analyze the subtypes of helper T (Th) cells among peripheral blood mononuclear cells (PBMCs). The effect of ADAM10 on PBMCs was analyzed in vitro. RESULTS: The levels of inflammatory cytokines and production of pathogenic Th cells were found to be higher in Korean patients with Buerger's disease. IL-17 treatment induced HMGB1 associated molecules. HMGB1 also induced IL-17 and Th17 associated transcription factors in Buerger's patients. We observed that ADAM10 regulates the interaction between IL-17 and HMGB1 via advanced glycation end products (RAGE)/nuclear factor-kappa B (NF-kB) pathway in patients with Buerger's disease. CONCLUSIONS: This study suggests that IL-17 and HMGB1 cytokines contribute to the pathogenesis of Buerger's disease. These results indicate that ADAM10 alleviates inflammation in Buerger's disease via the HMGB1 and RAGE/NF-κB signaling pathway and provides insights into the molecular basis of and a potential therapeutic strategy for Buerger's disease.

Thromboangiitis Obliterans Biomarker Shifts in Different Acute Phase Stages: A Case Study.

Thromboangiitis obliterans (TAO) is a rare vasculopathy that is predominantly seen in young male smokers. Recently, new biomarkers have been shown to be useful in distinguishing TAO from acute phase TAO in an Asian study population. The present case study illustrates their application in a European patient during TAO exacerbation and their association with therapeutic performance.

Humoral and cellular immune response to Buerger's disease.

OBJECTIVE: Thromboangiitis obliterans is a nonatherosclerotic occlusive disease, affecting small to moderate sized arteries of the upper and lower extremities, leading to progressive inflammation and clot formation. However, the role of humoral and cell-mediated immunity in the development of this disease has not been clearly identified. The present study was intended to investigate the humoral and cellular immune response in patients with Buerger's disease with different disease severity. METHODS: In an observational study, 80 male patients with Buerger's disease were included and categorized into three groups (mild, moderate, and severe) based on clinical manifestations. After blood sampling, cellular phenotypes were determined, and erythrocyte sedimentation rate, immunoglobulins (Ig) A, M, G, and E, as well as C3 and C4 components of the complement system and complement hemolytic activity (CH50) were measured. RESULTS: The mean age of the patient was 42.85 ± 8.39 years. Pulse abnormality, cold intolerance, and claudication were the most common symptoms. Eleven (13.75%), 46 (57.50%), and 23 (28.75%) patients had mild, moderate, and severe symptoms. Regression analyses showed that the presence of severe symptoms was significantly associated with elevated erythrocyte sedimentation rate and C4 levels (p < 0.05). CONCLUSION: Buerger's disease in severe cases was associated with increased erythrocyte sedimentation rate and abnormal C4 levels. The alterations in these inflammatory biomarkers might be due to a secondary inflammatory response to the presence of ulcer or gangrene and the inflammatory process in patients with severe symptoms.

Fibrinogen, Neutrophil-to-Lymphocyte Rate and Platelet-to-Neutrophil Rate as Novel Acute Phase Indicators in Patients with Thromboangiitis Obliterans.

BACKGROUND/OBJECTIVES: The acute exacerbations and progressive deterioration seen in thromboangiitis obliterans (TAO) have been related to poor clinical outcomes. Here, we have studied the association of laboratory biomarkers with the acute phase of TAO (AP-TAO). METHODS/RESULTS: We conducted a retrospective case-control study on 112 patients with TAO and 98 healthy controls; comparing the neutrophil-to-lymphocyte rate (NLR), lymphocyte-to-monocyte rate (LMR), platelet-to-neutrophil rate (PNR), fibrinogen (FIB), and apolipoprotein A-I (ApoA-I). Significantly higher NLR level, as well as lower LMR, PNR, and ApoA-I levels were observed in patients with TAO, particularly the acute phase. Significantly increased FIB was only observed in AP-TAO. A positive correlation was found between NLR and with C-reactive protein (CRP) in the acute phase (r = 0.817, P < 0.001). Moreover, NLR, PNR, and FIB levels of 3.38, 45.12, and 3.69 were shown to be the predictive cut-off values for the AP-TAO (sensitivity 72.5, 82,4, and 66,7%, specificity 92.2, 78.4, and 96.1%; area under the curve [AUC] 0.875, 0.855, and 0.872), respectively. The FIB level was independently associated with the AP-TAO (OR = 11.420, P = 0.007). CONCLUSIONS: NLR, PNR, and FIB may be useful markers for the identification of inflammation and the AP-TAO. FIB may be an independent risk factor for the acute phase.

Case report of two pregnancies and deliveries by a woman with Buerger disease.

Buerger disease is a chronic inflammatory disease that involves blood clot formation in the medium and small arteries, resulting in thrombophlebitis. It is usually observed in middle-aged men who smoke and is very rare in young women. Previous reports have indicated that Buerger disease worsens during pregnancy due to hypercoagulability associated with pregnancy, and newborns' birth weights were often lower than normal. This report describes a young woman with Buerger disease who experienced two pregnancies and deliveries. During the 1st pregnancy, d-dimer and soluble fibrin levels slightly increased, but no treatment was needed. However, during the 2nd pregnancy, d-dimer and soluble fibrin levels abruptly increased at 20 weeks of pregnancy, and heparin was administered subcutaneously. Four days after heparin administration, d-dimer and soluble fibrin levels decreased to normal pregnancy levels. d-dimer and soluble fibrin measurements were useful for evaluating the coagulation tendencies of this pregnant woman with Buerger disease.

D-dimer levels in patients with thromboangiitis obliterans.

Background: Thromboangiitis obliterans (TAO) or Buerger disease is a recurring progressive segmental vasculopathy that presents with inflammation and thrombosis of small and medium arteries and veins of the hands and feet. The exact cause remains unknown, with tobacco use (primarily smoking but also smokeless tobacco) being highly associated with the disease. The diagnosis is clinical and the lack of a diagnostic gold standard is a deterrent to diagnosing it in patients with atypical presentations. Obliterative endarteritis occurs perhaps due to a mixture of thrombosis and inflammation. The diagnostic sensitivity and specificity of D-dimer as a biomarker for thrombosis is well reported from its use in other areas such as deep vein thrombosis. Identification of a biomarker linked to the causation yields a diagnostic adjunct with a role in therapeutic decision-making, aiding diagnosis in atypical presentation, monitoring disease activity and gauging response to therapy. Methods: Between April 2014 and May 2015, we studied serum D-dimer (a marker of thrombosis) in 62 patients with TAO and compared this to 330 normal age- and sex-matched controls. We included all patients with peripheral arterial disease clinically diagnosed to have TAO according to the Shionoya criteria. There was no history of thrombosis or arterial disease in the control group. The control group was matched for baseline characteristics such as age and sex. All patients underwent a standard diagnostic protocol including blood tests (haemoglobin and creatinine), electrocardiogram, chest X-ray and ankle brachial pressure index. Blood was collected using an evacuated tube system into a citrate anticoagulant tube for testing D-dimer. Results: All the 62 patients diagnosed to have TAO were men with an average age of 40 years (range 18-65 years). They all had a history of tobacco use and did not have other atherogenic risk factors (part of the diagnostic criteria). Medium-vessel involvement was present in 53 patients (85%) and the rest presented with additional involvement of the popliteal and femoral vessels. Upper limb involvement or superficial thrombophlebitis was present in 95% of patients. Laboratory and imaging studies were consistent with TAO. The groups were well matched for age (p = 0.3). The median and interquartile range for D-dimer values were 61 ng/ml and 41-88 ng/ml in controls (n = 330) and 247 ng/ml and 126478 ng/ml in patients (n = 62), respectively (p<0.001). Conclusions: D-dimer levels are considerably elevated in patients with TAO. This indicates an underlying thrombotic process and suggests its potential role as a diagnostic adjunct. It also leads us to hypothesize a potential therapeutic benefit of anticoagulants in this disease.

Thromboangiitis obliterans episode: autoimmune flare-up or reinfection?

BACKGROUND: The possible role of infectious pathogens in the development of thromboangiitis obliterans (TAO) was considered soon after the disease was first described. However, it is not yet known whether infectious pathogens induce thrombotic vasculitis or if they cause a type of autoimmune disease. To investigate whether TAO relapses are more likely due to reinfection or autoimmune flare, the serum levels of toll-like receptor (sTLR) 4, sTLR2, C-reactive protein (CRP), and neopterin were evaluated in TAO patients during both the acute and quiescent phases of the disease as well as in a gender-, age-, and smoking habit-matched control group. METHODS: Following a cross-sectional study design, 28 patients in the acute phase of TAO and 23 patients in the quiescent phase participated in this study. In addition, 31 matched controls were enrolled. RESULTS: Toll-like receptor (TLR) 4 was significantly higher in patients in the acute phase of the disease than in patients in the quiescent phase (P=0.012). Also, TLR4 was significantly higher in the patients with CRP >7 µm/mL than in the patients with lower CRP (P=0.031). Notably, TLR4 in the patients in the quiescent phase of TAO was significantly lower than in the controls (P=0.006). No significant difference in the level of TLR2 was found among the groups (P>0.05). Neopterin was significantly higher in the acute phase of TAO in comparison to the quiescent phase (P=0.003) and the controls (P=0.005). CONCLUSION: These findings indicate that the trigger of TAO might be Gram-negative bacteria, which can be hidden or immunologically suppressed in the quiescent phase of TAO, leading to a lower level of TLR4 accompanying the normal level of neopterin. However, relapses might develop according to toxic or hypoxic cell injuries. Hence, TLR4 shedding will increase, and therefore, sTLR4 could become closer to the level demonstrated in the controls.

Alleviation of A disintegrin and metalloprotease 10 (ADAM10) on thromboangiitis obliterans involves the HMGB1/RAGE/ NF-κB pathway.

Thromboangiitis obliterans (TAO), also known as Buerger's disease, is a nonatherosclerotic inflammatory disease that influences medium- and small-sized blood vessels of extremities. However, mechanisms underlying TAO are still unclear. As a mediator associated with inflammation, A disintegrin and metalloprotease 10 (ADAM10) was hypothesized to play inhibitory roles in the development of TAO. Thus, the objective of this study is to investigate the effects of ADAM10 in a sodium laurate-induced TAO rat model and elucidate underlying mechanisms. Male Wistar rats were randomly divided into four groups (n = 6) for treatment: sham-operated (SHAM), TAO model (TAO), ADAM10 low dose injection (3 mg/kg; ADAM10-LD) and ADAM10 high dose injection (6 mg/kg; ADAM10-HD). After 14-day treatment, color Doppler ultrasound and hematology analysis indicated TAO rats displayed higher whole blood viscosity and blood platelet count compared with those in the SHAM group. Histologic evaluation and transmission electron microscopy revealed that the ultrastructural damages of vascular smooth muscle and endothelial cells were observed in TAO rats, such as fractured endoplasmic reticulum, decreased cell counts, and fibrillation. On the other hand, the typical signs and symptoms of TAO rats were significantly alleviated via ADAM10 treatment with a dose-dependent pattern. Real-time PCR and western blot results revealed that the expression of high-mobility-group box 1 (HMGB1), receptor for advanced glycation end-products (RAGE) and nuclear factor-kappa B (NF-κB) increased in TAO rats whereas decreased by ADAM10 treatment in both mRNA and protein levels. In conclusion, the results suggest ADAM10 alleviates symptoms of sodium laurate-induced TAO in rats via the RAGE/NF-κB signaling pathway and provides insight into the molecular basis and a potential therapeutic strategy for TAO.

Investigation of Asymmetric and Symmetric Dimethylarginine Levels after Iloprost Treatment in Patients with Buerger's Disease.

OBJECTIVE: The aim of this study was to compare the levels of acetyl-dimethylarginine (ADMA), symmetric dimethylarginine (SDMA), and the l-arginine/ADMA ratio before and after iloprost treatment in patients with Buerger's disease. METHODS: Between January 2011 and December 2015, data from 44 patients (36 males, 8 females, mean age 48.7 ± 18.1 years) with the diagnosis of Fontaine Stage III-IV Buerger's disease were included. Iloprost infusion was administered intravenously through the forearm veins for 7 days at a dose of 0.5-1.5 ng/kg/min over 16 h. Blood samples were collected before and after treatment for measurement of ADMA, SDMA, and l-arginine. ADMA, SDMA, l-arginine levels were measured using high performance liquid chromatography (HPLC). RESULTS: After iloprost treatment, ADMA and SDMA levels significantly decreased (p = .001). The increase in the l-arginine levels was not significant (p = .16). However, the l-arginine/ADMA ratio increased significantly (p = .001). CONCLUSION: Iloprost treatment decreases ADMA and SDMA, which are associated with endothelial dysfunctions in patients with Buerger's disease. Of note, the still higher than normal range of SDMA levels after iloprost treatment suggests that treatment should continue until SDMA levels are within the normal range in this patient population.

Rickettsia infection could be the missing piece of the Buerger's disease puzzle.

BACKGROUND: Rickettsia was suggested as a possible etiology of Buerger's disease (BD) in the 1980s but this suggestion was never ruled out or proven. Recently, we found evidence of Rickettsia by polymerase chain reaction in 3 out of 25 biopsy samples from the amputated limb of a young man diagnosed with BD. The aim of this paper was to investigate the presence of anti-rickettsial antibodies in the sera of BD patients. METHODS: To detect the IgG class antibody against Rickettsia rickettsii, which has cross reactions with the spotted fever group (RSFG), and Rickettsia typhi, which has cross reactions with typhus fever group, the sera of patients and controls were diluted to 1:64 and analyzed by indirect micro fluorescence immunoassay (MIF). RESULTS: The MIF study showed that 26 of the 28 patients were positive for Rickettsia rickettsii antibodies and MIF had the same appearance as the positive control, which was provided with the kit. In all members of the healthy control group, Rickettsia rickettsii was negative and had the appearance of the negative control. Rickettsia typhi was negative for all patients and members of the control group. CONCLUSIONS: A species of Rickettsia associated with the RSFG, which might not be pathogenic for the entire population, may induce BD in the context of a specific genetic or environmental background. RSFG infection could explain key questions about BD, including its gender and geographical distribution, clinical manifestation, angiography pattern, and pathological findings. Evaluating antibodies against RSFG in BD patients from different countries is now highly recommended.

Increased Serum HMGB-1, ICAM-1 and Metalloproteinase-9 Levels in Buerger's Patients.

BACKGROUND: Thromboangiitis obliterans (TAO), or Buerger's disease, is an inflammatory occlusive disorder that affects the limb arteries of young smokers. In the aetiology of TAO the immune system appears to play a critical role; however, information on the aspects involved in the evolution of vascular tissue inflammation and of this disease are still limited. OBJECTIVE: This study was carried out to investigate HMGB-1 (high mobility group box-1), MMP (matrix metalloproteinase)- 2, MMP-9, MMP-11 and ICAM (intercellular adhesion molecule)-1 circulating levels in subjects with Buerger's disease. METHODS: Between January 2010 and December 2012, eight patients underwent surgical revascularization of the lower limbs and a specimen of the affected arterial wall was obtained for histological confirmation of Buerger's disease. A blood sample was collected on the same day for measuring HMGB-1, MMP-3, MMP-9 and ICAM-1 by western blot analysis. Controls (n=7) were healthy non-smokers. RESULTS: TAO subjects had a significant increase in HMGB-1, MMP-9 and ICAM-1 compared with controls (P<.0001), while no differences were observed in MMP-2 and MMP-11 levels. Histology confirmed a strong inflammatory infiltrate with signs of necrosis in the arterial wall. CONCLUSION: These data suggest a role for HMGB -1 in the vascular lesions associated with TAO, unveiling HMGB-1 as a potential target for treating this rare disease.

Hypoxia-inducible factor-1α expression in the different stages of rat thromboangiitis obliterans.

We investigated the expression and effects of hypoxia-inducible factor-1α (HIF-1α) in rat thromboangiitis obliterans (TO). Rats were divided into sham and model groups. The model group was further divided into groups based on observation duration. Lauric acid was injected below an artery clamp to simulate TO in the model group; saline was used in the sham group. Clamps were removed 15 min after injection in both groups, and physiological changes were observed at different times (gross observation and hematoxylin and eosin staining). The animals were killed at various times following the operation and serum and muscle tissues were sampled. For the sham group: the endometrium was relatively intact; medial membrane and epineurium lesions were absent; and blood vessels and surrounding tissues had no inflammatory cell infiltration. For the model group: all subgroups displayed inflammation; large numbers of inflammatory cells were gathered; muscle tissue lost its normal texture and structure; and the internal elastic membrane was integrated. Compared with the preoperative status, HIF-1α expression increased significantly in all subgroups (P < 0.05); there was no change in the sham group. HIF-1α expression in each subgroup was different (F = 14.267, P < 0.05). Femoral artery injection of lauric acid can be used as a rat TO model owing to its simple application and success rate. HIF-1α expression increased in the early stage of TO and gradually decreased with the extension of ischemia time; it may play a leading role in TO development and can be used for diagnosis and cure evaluation.

Coexistence of axial spondyloarthritis and thromboangiitis obliterans in a young woman.

A peculiar coexistence of axial spondyloarthritis and ischemia of the feet and the fourth finger of the left hand in a young woman, who was a heavy smoker, is discussed in this report. This picture was considered within the context of thromboangiitis obliterans. Positivity of anti-nuclear antibodies and mild elevation of inflammatory parameters were noted. Computed tomography angiograms of upper and lower limbs showed luminal narrowing and occlusion of the left humeral, left anterior/posterior tibial and right anterior tibial arteries. Daily iloprost perfusions were started, and smoking cessation was strongly recommended. Coldness and rest pain in the distal extremities improved within a few weeks. The possibility that spondyloarthritis might precede the clinical picture of thromboangiitis obliterans should be considered in heavy smokers.

Noninvasive Transcutaneous Monitoring in Long-Term Follow-Up of Patients With Thromboangiitis Obliterans Treated With Intravenous Iloprost.

We evaluated the effectiveness of intravenous iloprost (IVI) in outpatients with thromboangiitis obliterans (TAO) and lower limb noninvasive transcutaneous monitoring (TCM) at follow-up (FU). Ten consecutive patients with TAO underwent IVI therapy. Transcutaneous oxygen (TcPo 2) and carbon dioxide (TcPco 2) determination and laser Doppler flowmetry (LDF) were performed before and after IVI at 3, 6, and 12 months of FU. Clinical response was positive in 7 patients, whereas 3 nonresponders underwent a second IVI cycle with 1 showing a late positive clinical response. After 12 months of FU, all patients were alive without amputations. Supine and dependent TcP2 levels significantly improved (P < .005). Hallux LDF values showed significant change with the maximal hyperemic test at 44°C (P < .005). Forefoot maximal hyperemic test at 44°C LDF (P < .005) and improved venous arterial reflex (P < .05) showed statistically significant time evolution. We demonstrated some degree of IVI effectiveness and evaluated TCM in patients with TAO.

G-protein coupled receptor auto-antibodies in thromboangiitis obliterans (Buerger's disease) and their removal by immunoadsorption.

BACKGROUND: Immunhistopathological and serological data favors an immunopathogenesis of thromboangiitis onliterans (TAO, Buerger's disease). Auto antbodies seem to play a major role. Immunoadsorption (IA) proved to be therapeutically effective. We focused on agonistic autoantibodies (agAAB) directed against G-protein coupled receptors (GPCR) and proved the hypothesis, that these agAAB might be present in TAO and that a five day course of IA might be able to eliminate these agAAB effectively. PATIENTS AND METHODS: Between December 2012 and May 2014 11 TAO-patients were treated by IA in a five day course. AgAAB-analysis was performed using specific ELISA techniques. RESULTS: AgAAB were detected in 9 out of 11 patients (81.8 %).Multiple agAAB were present in 7 patients (63.6 %). A clustering of agAAB directed against loop1 of the adrenergic α1-receptor and the endothelin-A-(ETA)receptor was identified, representing 72.7 % resp. 54.5 % of the patients. AgAAB directed against the angiotensin-1 (AT-1) epitope 1 or 2 were detected in 3 patients and agAAB directed against protease-activated receptor (PAR) loop1/2 were seen in 2 patients. AgAAB directed against ETA-receptor loop1 never appeared without agAAB directed against α1-receptor loop1. Immediately after a five day-course of IA agAAB were absent in 81.8 % of the total study group and in 77.8 % of all cases tested positive for agAAB before IA. CONCLUSIONS: AgAAB directed against GPCR were identified in TAO patients with a clustering of agAAB directed against α-1-adrenergic receptor loop1 and ETA-receptor loop1. AgAA were eliminated by IA in the majority of cases. We suggest that these agAA play an important role in the pathogenesis of TAO and that their elimination might be responsible for the positive therapeutic effects reported in patients treated with IA.

Sera of patients with thromboangiitis obliterans activated cultured human umbilical vein endothelial cells (HUVECs) and changed their adhesive properties.

INTRODUCTION: The aim of this study was to investigate the impact of thromboangiitis obliterans (TAO) sera on activation of primary cultures of human umbilical vein endothelial cells (HUVECs) as a model for vascular endothelial cells. METHODS: Study subjects included 21 TAO patients as the case group and 20 healthy smokers and 17 healthy non-smokers as control groups. Case and control groups were matched based on their age, socioeconomic status and smoking habit. HUVECs were incubated with the sera of case and control groups and gene expression of intercellular adhesion molecule (ICAM-1) and vascular adhesion molecule (VCAM-1) were evaluated by real-time polymerase chain reaction, TaqMan method. RESULTS: The expression of ICAM-1 and VCAM-1 were significantly higher in HUVECs after incubation with TAO sera compared to control groups (P < 0.05). VCAM-1 had a significant correlation with duration of smoking (P < 0.001, R = 0.672), while the expression of ICAM-1 had a significant correlation with the number of cigarettes smoked daily (P = 0.04, R = 0.421). CONCLUSION: Sera from TAO patients could activate HUVECs. This same activation might occur in vivo by the responsible cytokines, in particular those released from activated platelets, free oxygen radicals, and possibly low levels of nitric oxide (NO) of the sera of TAO patients, as a consequences of chronic cigarette smoking and of endothelial NO synthase polymorphism. Therefore, plasma exchange might be helpful in acute phase of the disease for saving the limbs and administration the combinations of exogenous NO with anti-oxidants might be helpful in long-term management of TAO patients to reduce the risk and rate of amputation.

Investigation of the expression of mediators of neovascularization from mononuclear leukocytes in thromboangiitis obliterans.

The aim of this study was to investigate the expression of the cytokines, chemokines and effective molecules of peripheral blood mononuclear cells (PBMCs) that play a role in neovascularization in thromboangiitis obliterans (TAO). Lymphocytes from TAO patients (n = 20) and control subjects (healthy smokers [n = 16] and non-smokers [n = 17]) were evaluated using realtime polymerase chain reaction in order to examine the mRNA expression of CXCL1 and interleukin 8 (IL-8; inducers of collateral development by recruitment of circulating progenitor cells [CPCs]), endothelial cell growth factor A (VEGF-A) and inducible nitric oxide synthase (iNOS; inducers of angiogenesis) and interferon gamma (IFN-γ) and vascular endothelial growth factor receptor 1 (VEGFR-1; inhibitors of angiogenesis). CXCL1 expression was significantly higher in the TAO patients than control subjects. The expressions of IL-8, VEGFR-1 and IFN-γ were significantly higher in the TAO patients and smokers than in non-smokers. However, no differences in iNOS and VEGF-A expression were noted. In conclusion, PBMCs from TAO patients expressed cytokines that potentially recruit CPCs and promote arteriogenesis. However, TAO patients typically have low CPC levels, perhaps due to high oxidative stress. Further studies are recommended in order to investigate the efficacy of antioxidant therapy on the outcome of TAO before administration of angiogenic factors.