Comparative Analysis of Transcriptome Profiles in Patients with Thromboangiitis Obliterans.

BACKGROUND: Thromboangiitis obliterans (TAO) causes vascular insufficiency due to chronic inflammation and abrupt thrombosis of the medium and small arteries of the extremities. In our study, we aimed to determine biomarkers for the diagnosis of TAO by evaluating 15 male TAO patients with Shinoya diagnostic criteria and 5 healthy controls who did not have TAO-related symptoms in their family histories. METHODS: The Clariom D Affymetrix platform was used to conduct microarray analysis on total RNA extracted from whole blood. A total of 477 genes (FC ≤ 5 or >5) common to the fifteen patient and five control samples were selected using comparative microarray analysis; among them, 79 genes were upregulated and 398 genes were downregulated. RESULTS: According to FC ≤ 10 or >10, in the same TAO patient and control group, 13 genes out of 28 were upregulated, whereas 15 genes were downregulated. The 11 key genes identified according to their mean log2FC values were PLP2, RPL27A, CCL4, FMNL1, EGR1, EIF4A1, RPL9, LAMP2, RNF149, EIF4G2, and DGKZ. The genes were ranked according to their relative expression as follows: FMNL1 > RNF149 > RPL27A > EIF4G2 > EIF4A1 > LAMP2 > EGR1 > PLP2 > DGKZ > RPL9 > CCL4. Using protein-protein interaction network analysis, RPL9, RPL27A, and RPL32 were found to be closely related to EIF4G2 and EIF4A1. The Reactome pathway found pathways linked to 28 genes. These pathways included the immune system, cellular responses to stress, cytokine signaling in the immune system, and signaling by ROBO receptors. CONCLUSIONS: By figuring out the protein expression levels of the genes that have been found to explain how TAO disease works at the molecular level, it will be possible to figure out how well these chosen transcripts can diagnose and predict the disease.

Integrated analysis of long non-coding RNA-microRNA-mRNA competing endogenous RNAregulatory networks in thromboangiitis obliterans.

Thromboangiitis obliterans (TAO) is a non-atherosclerotic, segmental, chronic vascular inflammatory disease. Our aim was to explore the underlying mechanisms of long non-coding RNA (lncRNA)-related competing endogenous RNAs (ceRNAs) in TAO. Six blood samples were collected from patients with TAO and healthy individuals (three for each category). Total RNA was extracted from the blood of each participant and sequenced. Differentially expressed lncRNAs (DE-lncRNAs) and miRNAs (DE-miRNAs) were screened, and ceRNA networks associated with TAO were constructed. Thereafter, the genes in the ceRNA network were subjected to functional analyses. Finally, a ceRNA relationship (lncRNA NEAT1-hsa-miR-1-3p-mRNA GNA12) was selected for further validation. Analysis revealed that 347 DE-lncRNAs (150 downregulated and 197 upregulated) and 16 DE-miRNAs (3 downregulated and 13 upregulated) were identified in TAO. Further, TAO-associated ceRNA networks, which included 219 lncRNAs, 6 miRNAs, and 53 mRNAs, were proposed and subjected to gene annotation and pathway analysis. Additionally, NEAT1 and GNA12 levels were significantly upregulated, while miR-1-3p levels were evidently downregulated in TAO patients, as compared with those in healthy controls. Dual luciferase reporter assays showed that NEAT1, miR-1-3p, and GNA12 interacted with each other. We report potential TAO-associated ceRNA regulatory networks and suggest activation of NEAT1/miR-1-3p/GNA12 signaling as a novel mechanism for TAO progression.

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.

Human leukocyte antigen class I (A, B) and class II (DRB1) allele and haplotype frequencies in Iranian patients with Buerger's disease.

OBJECTIVE: The aim of this study was to investigate the human leukocyte antigen (HLA) class I (HLA-A and HLA-B) and II (HLA-DRB1) allele and haplotype frequencies in a group of Iranian patients with Buerger's disease (BD) in comparison with a normal healthy control group. METHODS: A total of 70 unrelated male patients and 100 healthy controls from Sina Hospital, Tehran, Iran, belonging to the same ethnic background, were enrolled in this case-control study. HLA-A, B, and DRB1 typing were performed by polymerase chain reaction with sequence-specific primers (PCR-SSP). RESULTS: The results of this case-control study showed that the frequency of the HLA-A*03:01 (odds ratio (OR) = 2.88, P value (Pv) = .002), HLA-A*29:01 (OR = 15.31, Pv < .001), HLA-DRB1*04:02 (OR = 3.41, Pv < .001), and HLA-DRB1*16:01 (OR = 8.16, Pv < .001) was significantly higher in BD patients compared with healthy controls, whereas the frequency of the HLA-DRB1*01:01 (OR = 0.03, Pv < .001) was significantly lower in BD patients. The most frequent extended haplotypes in our patients were HLA-A*02:01-B*55:01-DRB1*04:03. CONCLUSION: This study is the first study evaluating an association between the HLA pattern and BD in the patients with BD from North West and North Iran.

[Comprehensive treatment of a patient with Buerger's disease using genetically engineered complexes VEGF-165]. / Kompleksnoe lechenie patsienta s bolezn'iu Biurgera s primeneniem genno-inzhenernykh konstruktsii VEGF-165.

Buerger's disease, also known as thromboangiitis obliterans, is a severe invalidating systemic vascular disease. To one of the modern methods, which is distinguished by its radically new principles of action, as well as holding much promise for further study and application in treatment of patients with lower limb chronic ischaemia induced by thromboangiitis obliterans belongs the use of genetically engineered complexes based on vascular endothelial growth factor VEGF-165 ('Neovasculgen'). 'Neovasculgen' is a genetically engineered complex being a circular DNA (native plasmid on the CELO vector and Ad5), carrying the human VEGF-165 gene, encoding VEGF synthesis. Injection of this drug to the ischaemised tissues of lower extremities ensures long-term synthesis of vascular endothelial growth factor 165 leading to the development of an additional collateral vascular network and consequently to increased perfusion of tissues with oxygen and decreased degree of ischaemia. Presented herein is a clinical case report of a successful therapeutic outcome achieved in a patient suffering for a long time from thromboangiitis obliterans (Buerger's disease) and treated with genetically engineered complexes based on vascular endothelial growth factor ('Neovasculgen') used as a component of comprehensive conservative therapy.

Pathogenesis of thromboangiitis obliterans: Gene polymorphism and immunoregulation of human vascular endothelial cells.

Thromboangiitis obliterans (TAO) is a nonatherosclerotic, segmental, inflammatory vasculitis, which commonly affects the small- and medium-sized arteries of the upper and lower extremities. Despite its discovery more than a century ago, little progress has been made in its treatment. Unless the pathogenesis is elucidated, therapeutic approaches will be limited. The purpose of this review article is to collate current knowledge of mechanisms for the pathogenesis of thromboangiitis obliterans and to propose potential mechanisms from a genetic and immunoreactive point of view for its inception. Therefore, we discuss the possibility that the pathogenesis of this disease is due to a type of gene polymorphism, which leads to an immunological inflammatory vasculitis associated with tobacco abuse, highly linked to T cells, human vascular endothelial cells (HVECs), and the TLR-MyD88-NFκB pathway, distinct from arteriosclerosis obliterans and other vasculitides.

Association Between Three SNPs and Thromboangiitis Obliterans in Xinjiang Uyghur Population.

BACKGROUND: Thromboangiitis obliterans (TAO), also called Buerger's disease, is a chronic peripheral vascular occlusive disease. It is an obliterative vasculitis characterized by arterial thrombosis and strongly associated with tobacco exposure. The pathogenesis and etiology of TAO are not well understood, but genetic factors may be important in its development. A case-control study was undertaken to identify genetic factors potentially involved in the pathogenesis of TAO in a Xinjiang Uyghur population of China, where TAO is common. METHODS: We ascertained 177 TAO patients by clinical screening and 86 healthy individuals from the HAPMAP database. The genotypes of single-nucleotide polymorphisms (SNPs) of the participants were identified using the Affymetrix Genome-Wide Human SNP Array 6.0 to perform a genome wide association study (GWAS). The association between the SNPs and incidence of TAO was quantified using race stratification exposure. RESULTS: Through a case-control GWAS study 26 SNPs were significantly associated with incidence of TAO following a Bonferroni correction. However, after genomic control correction for population stratification only three of these SNPS were highly significantly associated with TAO: rs376511 in IL17RC (OR = 24.4, 95% CI:8.68 - 68.62, p < 0.0001), rs7632505 in SEMA5B (OR = 29.47, 95% CI:7.16 - 121.3, p < 0.0001), and rs10178082 (OR = 18.09, 95% CI: 6.56 - 49.92, p < 0.0001) showed a significant risk of TAO in the Uyghur population. CONCLUSIONS: This study shows an association between these 3 SNPs and susceptibility to TAO in the Uyghur population, suggesting that polymorphisms in the IL-17RC and Sema 5B genes may pre-dispose individuals in this population to development of TAO. These findings require replication.

Association of the five gene related endothelial cell dysfunction polymorphisms with Buerger's disease development.

BACKGROUND: The aim of this study was to evaluate the impact of the polymorphisms of four genes related to vascular endothelium dysfunction on the development and outcome of Buerger's disease (BD). The genes studied were eNOS-786 T>C, eNOS894 G>T, ET-1 8000 T>C, PAI-1 4G/5G and ACE I/D. METHODS: Polymerase chain reaction and restriction fragment analysis were used to detect eNOS-786 T>C, eNOS894 G>T, ET-1 8000 T>C, PAI-1(4G/5G) and ACE(I/D) polymorphisms in 36 BD patients and 36 healthy individuals matched for race, age and gender. A decision tree for predicting BD was drawn using Rapidminer 5.3 software. RESULTS: The frequency of eNOS-T786C alleles was significantly different between the BD group and the healthy controls (P<0.001, OR:6.1). The frequency of PAI-1(4G/5G) alleles was significantly different between the BD group and the healthy controls (P=0.005, OR:4.9). The frequency of eNOS G894T alleles was not statistically different between BD and the healthy controls (P=0.09). No significant difference between allele frequency of ACE(I/D) was found (P=0.07). There was, also, no significant difference between the allele frequency of ET-1 8000 T>C (P=0.1). In logistic regression analysis, the C allele for eNOS-786 and 4G/4G for PAI-1 were significant for predicting BD. According to the decision tree, the proportion of the current gene-polymorphisms likely to develop BD was calculated as maximum 27.7%. CONCLUSIONS: It seems that eNOS-T786C, PAI-1(4G/5G) are important polymorphisms in developing BD. However, the decision tree might give confidence to the families of BD patients that if they maintain a healthy lifestyle, they may not develop BD.

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.

Embolus-carried vascular endothelial cell growth factor 165 improves angiogenesis in thromboangiitis obliterans.

We investigated neovasculization effects of embolus-carried human vascular endothelial cell growth factor 165 (VEGF165)-encoded adenovirus (Ad) vector in the hindlimbs of rats with thromboangiitis obliterans (TAO). Rats were equally divided into blank control (I), TAO model (II), embolus (III), Ad-VEGF165 intravascular treatment (IV), Ad-VEGF165 intramuscular treatment (V), and embolus-carried Ad-VEGF165 (VI) groups. After interventional treatment, the neovasculization effect of the test gene was observed using immunohistochemistry. At 1 week after administration, compared with group II, groups V and VI had significantly increased microvessel densities, but no significant difference was observed between groups V and VI. At 2 weeks, groups V and VI exhibited significantly increased microvessel densities. At 1 week after administration, compared with group II, both groups V and VI showed a significant difference in the ratio between the α-smooth muscle actin count and the muscle fiber count, whereas no significant difference was observed between them. At 2 weeks, groups V and VI also exhibited significant differences in these ratios compared with the other groups. We conclude that Ad-VEGF165 promotes neovasculization in ischemic limbs. Embolus-carried Ad- VEGF165 had the most pronounced effect.

Possible involvement of Notch signaling in the pathogenesis of Buerger's disease.

PURPOSE: Under pathological conditions, the Notch signal pathway is involved in the inflammatory process in arteriosclerosis, atherosclerosis and angiogenesis under ischemic conditions. The purpose of this study was to observe whether or not Buerger's disease is associated with Notch signal activation. METHODS: All the patients were diagnosed between 1980 and 2009 at Nagoya University Hospital. Twenty-two specimens from 12 patients with Buerger's disease (TAO) and 13 specimens from nine patients with arteriosclerosis obliterans (ASO) were analyzed by immunohistochemistry for Notch1, Jagged-1 (a Notch ligand) and Hes-1 (a Notch 1 target transcription factor). RESULTS: Notch1 and Jagged-1 were highly expressed in the endothelium in the new vasa vasorum and in the smooth muscle cells in the media of specimens from both groups. These Notch-related proteins were also remarkably expressed in inflammatory cells in the intima of specimens from TAO patients. Fewer inflammatory cells expressed Notch-related proteins in atheromatous plaques (Notch1 (%): 8.4 ± 0.76 versus 1.3 ± 0.43, P < 0.001; Jagged-1(%): 9.3 ± 1.1 versus 5.2 ± 1.1, P = 0.03). Indeed, Hes-1, which is a transcription factor downstream of Notch1, was remarkably expressed in the endothelium of new capillary vessels and inflammatory cells in TAO patients. Notch1-positive mononuclear cells were also seen in the thrombus in samples from the TAO group. CONCLUSIONS: Our findings are the first demonstration that Notch signal activation in inflammatory cells may be involved in the pathophysiological mechanism underlying Buerger's disease.

Novel risk factors for premature peripheral arterial occlusive disease in non-diabetic patients: a case-control study.

BACKGROUND: This study aimed to determine the prevalence of genetic and environmental vascular risk factors in non diabetic patients with premature peripheral arterial disease, either peripheral arterial occlusive disease or thromboangiitis obliterans, the two main entities of peripheral arterial disease, and to established whether some of them are specifically associated with one or another of the premature peripheral arterial disease subgroups. METHODS AND RESULTS: This study included 113 non diabetic patients with premature peripheral arterial disease (diagnosis <45-year old) presenting either a peripheral arterial occlusive disease (N = 64) or a thromboangiitis obliterans (N = 49), and 241 controls matched for age and gender. Both patient groups demonstrated common traits including cigarette smoking, low physical activity, decreased levels of HDL-cholesterol, apolipoprotein A-I, pyridoxal 5'-phosphate (active form of B6 vitamin) and zinc. Premature peripheral arterial occlusive disease was characterized by the presence of a family history of peripheral arterial and carotid artery diseases (OR 2.3 and 5.8 respectively, 95% CI), high lipoprotein (a) levels above 300 mg/L (OR 2.3, 95% CI), the presence of the factor V Leiden (OR 5.1, 95% CI) and the glycoprotein Ia(807T,837T,873A) allele (OR 2.3, 95% CI). In thromboangiitis obliterans group, more patients were regular consumers of cannabis (OR 3.5, 95% CI) and higher levels in plasma copper has been shown (OR 6.5, 95% CI). CONCLUSIONS: According to our results from a non exhaustive list of study parameters, we might hypothesize for 1) a genetic basis for premature peripheral arterial occlusive disease development and 2) the prevalence of environmental factors in the development of thromboangiitis obliterans (tobacco and cannabis). Moreover, for the first time, we demonstrated that the 807T/837T/873A allele of platelet glycoprotein Ia may confer an additional risk for development of peripheral atherosclerosis in premature peripheral arterial occlusive disease.

A single nucleotide polymorphism in the 3'-untranslated region of MyD88 gene is associated with Buerger disease but not with Takayasu arteritis in Japanese.

Buerger disease (BD) and Takayasu arteritis (TA) are rare vascular disorders. Although their etiology and pathogenesis have not been elucidated, several studies have suggested the involvement of innate immunity. Myeloid differentiation primary-response protein 88 (MyD88) is a key signaling adaptor for all Toll-like receptors, which have a central role in innate immunity. In the present study, we evaluated the association of MyD88 with BD and TA. We conducted case-control studies in Japanese populations composing of 131 BD cases, 90 TA cases and 270 healthy controls to be genotyped for a single nucleotide polymorphism rs7744 A>G in the 3'-untranslated region of MyD88 gene. The frequency of GG genotype was significantly lower in the BD patients than in the controls (6.9 vs 15.9%, P=0.011, odds ratio=0.39, 95% confidence interval; 0.19, 0.81), although there was no significant difference in the genotype frequencies between the TA patients and controls. It was suggested that MyD88 may confer resistance to BD in Japanese. Because this is the first report of the association between MyD88 and BD, replication studies in other cohorts are required.

Transfection of human HGF plasmid DNA improves limb salvage in Buerger's disease patients with critical limb ischemia.

AIM: Hepatocyte growth factor is a potent angiogenic agent. This study investigated the efficacy and safety of intramuscular injection of naked plasmid DNA encoding the human hepatocyte growth factor gene in Japanese patients with Buerger's disease and critical limb ischemia. METHODS: An open-label clinical study was performed at eight hospitals in Japan from May 2004 to April 2008. Ten patients were enrolled. They had Buerger's disease with ischemic ulcers, were not candidates for revascularization, and were unresponsive to conventional drug therapy. Treatment consisted of 8 injections (total dose: 4 mg) of hepatocyte growth factor plasmid, which were administered into the calf muscles and/or distal thigh muscles of the ischemic limbs under ultrasound guidance. Administration was done twice at an interval of 4 weeks. If there was no improvement after 2 doses, a 3rd dose could be administered. The response to treatment was evaluated from the reduction of ischemic ulcer size. RESULTS: The size of ischemic ulcers showed a decrease in 6/9 (66.7%) patients and the ulcers healed completely in 5/9 (55.6%) patients after gene therapy. Major amputation was not required. There were no deaths and no major safety concerns. CONCLUSION: Hepatocyte growth factor gene therapy is safe and effective for critical limb ischemia in patients with Buerger's disease.

Effect of eNOS and ET-1 polymorphisms in thromboangiitis obliterans.

Thromboangiitis obliterans (TAO), or Buerger disease, is a segmental occlusive inflammatory disorder of the arteries and veins, and etiopathogenesis is still obscure. It is strongly connected to the use of tobacco products, especially smoking. Smoking cessation is obligatory for success of the medical treatment. In the current study, we investigated the prevalence of endothelial nitric oxide synthase (eNOS) 894 G-->T and endothelin-1 (ET-1) 8000 T-->C polymorphisms in association with TAO to reveal any possible involvement in the TAO pathophysiology. The T allele of the eNOS 894 G-->T polymorphism was found to be associated with the prevention of TAO.

Apolipoprotein C1 and apolipoprotein E are differentially expressed in atheroma of the carotid and femoral artery.

BACKGROUND: A number of the genes and proteins as the causes of carotid atherosclerotic disease have been recently reported, but the major factors for atherosclerosis have still not been identified. METHODS: The atherosclerotic atheromas were obtained during endarterectomy for each of 10 cases of diseased carotid and femoral arteries. As the nonatherosclerotic arteries, the iliac arteries were obtained during organ harvest from five cases of brain-dead donors, and the leg arteries were obtained during leg amputation from five cases of Buerger's disease. The total RNAs and proteins were isolated from the atheromas and arteries. The annealing control primer method was used to screen the differentially expressed mRNAs. To identify if the mRNA expression of screened gene was associated with the protein expression, we performed an immunohistochemical analysis. RESULTS: We found that the apolipoprotein C1 (apo C1) gene was prominently expressed in the atheroma of the carotid and femoral arteries, as compared to the nonatherosclerotic arteries. Immunohistochemical analysis showed the high expression of apo C1 protein in the atheromas of the carotid and femoral arteries. Apo E protein was also highly expressed in atheromas compared with the nonatherosclerotic arteries, but there was no difference for apo C2 protein between those four groups of arteries. DISCUSSION: The expression of apo C1 and apo E are closely associated with the susceptibility to the pathogenesis of atherosclerosis. This study suggests that these factors might play important roles in the future to screen for preventing atherosclerosis and for diagnostic testing of patients.

Synergistic contribution of CD14 and HLA loci in the susceptibility to Buerger disease.

Buerger disease (BD) is an occulusive vascular disease of unknown etiology. Although cigarette smoking is a well-known risk factor of BD, genetic factors may also play a role in the etiology. Because chronic bacterial infection such as oral periodontitis is suggested to be involved in the pathogenesis of BD, gene polymorphisms involved in the infectious immunity might be associated with BD as the genetic factor(s). We have previously reported that HLA-DRB1*1501 and B54 was associated with BD in Japanese. In this study, polymorphisms in HLA-DPB1, DRB1 and B were analyzed in 131 Japanese BD patients and 227 healthy controls. In addition, we investigated a functional promoter polymorphism, -260 C > T, of CD14 that is a main receptor of bacterial lipopolysaccharide. It was found that the frequencies of CD14 TT genotype [37.4 vs. 24.2%, P = 0.008 OR = 1.87, 95% confidence interval (CI); 1.18, 2.97], DRB1*1501 (34.4 vs. 13.2%, P (c) = 4.4 x 10(-5), OR = 3.44, 95%CI; 2.06, 5.73) and DPB1*0501 (79.4 vs. 55.1%, P (c) = 4.7 x 10(-5), OR = 3.14, 95%CI; 1.93, 5.11) were significantly higher in the patients than in the controls, demonstrating that at least three genetic markers were associated with BD. Stratification analyses of these associated markers suggested synergistic roles of the genetic factors. Odds ratios ranged from 4.72 to 12.57 in individuals carrying any two of these three markers. These findings suggested that the susceptibility to BD was in part controlled by genes involved in the innate and adaptive immunity.

Platelet receptor HPA-1 polymorphism of alphaIIbbeta3 and 807 C/T polymorphism of alpha2beta1 and Buerger's disease.

Thromboangiitis obliterans or Buerger's disease is an episodic and segmental inflammatory and thrombotic process of the medium and small arteries of the lower extremities. Even though the disease was described 90 years ago, the etiopathogenesis is still under consideration. Afflicted patients are mostly young male cigarette smokers without signs of atherosclerosis or other risk factors for peripheral arterial occlusive disease. This indicates that hereditary thrombophilic factors could play a role in the etiopathogenesis. Recently, increasing evidence shows that platelet receptor polymorphisms (HPA-1 polymorphism of beta3 subunit of alphaIIbbeta3 and 807 C/T polymorphism alpha2beta1) are associated with early onset of arterial thrombosis (myocardial infarction, stroke). This case-control study was designed to assess whether the 807 C/T polymorphism or the HPA-1 polymorphism is involved in the pathogenesis of Buerger's disease or has any influence on the clinical course of Buerger's disease. Eighteen patients with Buerger's disease and 81 (sex and age matched) healthy control subjects (mean age 44 +/- 10 vs 45 +/- 8 years, respectively) were genotyped for platelet receptor HPA-1 and GPIa 807 C/T polymorphism. The gene frequency of HPA-1 and GPIa 807 C/T polymorphisms was identical in both groups. Prevalence of hetero- and homozygous carriers of the HPA-1b allel (1a1b and 1b1b genotype) as well as the prevalence of the 807 C/T and 807 T/T carriers did not differ significantly between the two groups, p >0.05. The grade of clinical disease manifestation as well as disease progression did not reveal any significant relationship with HPA-1 and 807 C/T polymorphisms. A relationship between the age at onset of the disease and HPA-1 polymorphism was not found. Otherwise analysis of the GPIa 807 C/T platelet receptor polymorphism showed that the average age of patients who are carriers of the T allele at early onset of disease was 32 +/- 6 years (range 27-48 years) compared to 42 +/- 6 years (range 34-53 years) of the C/C carriers (p <0.05). This indicates that the GPIa 807 C/T polymorphism does not represent a risk factor for Buerger's disease itself, but could be associated with premature onset of this disorder in predisposed individuals.