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Funding Acknowledgements Type of funding sources: Other. Main funding source(s): Private grant from Kowa Pharmaceuticals to Brigham and Woman's Hospital Calcific aortic valve stenosis (CAVS) is the most prevent valvular heart disease in the western world increasing exponentially with age, with an 112% increase in CAVS deaths in the last three decades; however no therapeutic treatment is currently available. Recently, lipoprotein(a) [Lp(a)] has been demonstrated to be an independent and causal risk factor for CAVS, yet the understanding of its cellular uptake and catabolism is limited thus underscoring the need for further investigation. This study aimed to determine a target receptor, unique for Lp(a) on the surface of valvular interstitial cells (VICs) and ascertain the role of the receptor on the development of VIC calcification. Unbiased ligand-receptor capture mass spectrometry (TriCEPS) was used to identify target receptor, with western blotting, ELISA, qPCR, alizarin red calcium staining and immunofluorescence used to validate the targets in vitro via siRNA inhibition and overexpression. Transmission electron microscopy (TEM) was used to determine uptake of Lp(a) within excised human valves. Identification of small molecule inhibitors was assessed computationally via the L1000 dataset, with the top hit candidate validated in vitro. Genotype-phenotype studies were examined using the United Kingdom Biobank (UKBB) and the Millions Veterans Program. Linear regression was used to evaluate association between aortic stenosis and plasma Lp(a) levels, and a phenotype-wide association analysis was then performed against this generated 'genotype'. Ligand-receptor capture mass spectrometry was used to detect novel membrane proteins with specific binding to Lp(a); MFSD5, MRC2, LDLR were identified as possible candidates. MFSD5 RNAscope demonstrated its presence in human aortic valves. Lp(a) uptake in VICs was confirmed via western blot and TEM. MFSD5 siRNA significantly reduced dil-labelled Lp(a) uptake in human VICs (p=0.003) and HEPG2 cells (p=0.0003), conversely MFSD5 overexpression increased uptake (p=0.0345, p=0.0318), whilst specificity of MFSD5 to Lp(a) alone was shown via no change in LDL uptake following MFSD5 inhibition (p=0.616, p=0.991). MFSD5 inhibition reduced RUNX2 (p=0.0124) and Osteocalcin (p<0.001) RNA expression and reduced alizarin red staining following culture in Lp(a) osteogenic media for 21 days (p<0.0033). Druggability of MFSD5 was confirmed by the L1000 database, which identified aminopurvalanol as a binding partner for MFSD5 and significantly reduced Lp(a) uptake within VICs (p=0.0091). MFSD5-loss of function within the UKBB showed no significant cardiovascular association, however 50kb +/- of the MFSD5 gene showed nominal association with hyperlipidaemia and atrial fibrillation. The current study demonstrates the novel Lp(a) receptor MFSD5 may be responsible for uptake of Lp(a) within VICs, resulting in the development of aortic valve calcification, highlighting the need for further exploration into the role of MFSD5 in aortic valve disease. [ABSTRACT FROM AUTHOR]

Early pancreatic cancer is small and limited to the pancreas. In contrast, small pancreatic cancer may include peripancreatic vasculature or metastasis involvement. This study evaluates images of early pancreatic cancer on multidetector CT (MDCT) using contrast-enhanced multiphasic imaging, and post-processed pancreatic duct images. CT findings and pathological features were analysed in eight patients with early pancreatic cancer. Pathological evaluation included location, size and histological grading of the tumour. MDCT evaluation covered the maximum diameter of the main pancreatic duct (MPD), stenosis or obstruction of the MPD, loss of normal lobar texture and associated pancreatitis. Attenuation differences between normal pancreatic parenchyma and the tumour (AD-PT) were also measured. Focal stenosis or obstruction of the MPD with dilatation of the distal MPD was demonstrated in all patients. Associated pancreatitis occurred in six patients with tumours measuring 12 mm or greater. Loss of normal lobar texture was recognised in four cases with the tumour measuring 14 mm or greater. Statistically, low-attenuated lesions and high-attenuated lesions differed with respect to the tumour size (p,0.01), and a positive relationship was demonstrated between the tumour size and AD-PT (r50.84). In seven cases, AD-PT is higher during the arterial phase than the pancreatic phase. Early pancreatic cancer appears as low attenuation on early phase, and as high- to iso-attenuation during the pancreatic and delayed phases in respect to the tumour size. Focal stenosis or obstruction of the MPD with dilatation of the distal MPD observed on curved reformation imaging seems important in the diagnosis of early pancreatic cancer. [ABSTRACT FROM AUTHOR]

X-linked agammaglobulinaemia (XLA) is an inherited immunodeficiency that is caused by a block in early B-cell differentiation. Whereas early B precursors in the bone marrow are present in substantial numbers, XLA-affected individuals have dramatically reduced numbers of circulating mature B cells, plasma cells and immunoglobulins of all isotypes. We report on a Japanese family with 3 XLA patients, in whom the serum immunoglobulin levels and number of B cells showed a significant difference among them in spite of harbouring the same splice donor site mutation in the BTK gene. We developed concise method for detection of this mutation, which is helpful for discovering the carrier. Patient 2 showed a significant serum immunoglobulin levels of all isotypes, including allergen-specific IgE. Expression of a normal and truncated size BTK gene was detected in patient 2′s peripheral blood mononuclear cells (PBMCs). Expression of BTK protein was also detected in some B cells. These results suggest that the leaky phenotype in patient 2 was caused in part by the expression of a normal BTK gene transcript. The increased frequency of infection with age expanded the number of B cells with normal BTK gene expression and produced the serum immunoglobulin, including IgE. [ABSTRACT FROM AUTHOR]

The alanine-rich repeating region (A-region) in the surface protein antigen (PAc) of Streptococcus mutans has received much attention as an antigenic component for vaccines against dental caries. The PAc (residue 361–386) peptide in the A-region possesses a multiple binding motif (L- -V-K- -A) to various HLA-DR molecules and a B-cell core epitope (- Y- - -L- -Y- - - -) that recognizes the inhibiting antibody to S. mutans. In the present study, we investigated the immunogenicity of the PAc (361–386) peptide in humans and regulators of induction of the anti-PAc (361–386) peptide IgA antibody (aPPA) in saliva. The PAc (361–386) peptide was confirmed as an ideal peptide antigen for induction of the inhibiting antibody to S. mutans in 151 healthy human subjects (36·6 ± 12·6 years old) by quantitative analyses of oral bacteria and ELISA, as the aPPA titre in human saliva decreased significantly in an age-dependent manner. Homozygous DRB1* 0405 and 1502, and heterozygous DRB1* 0405/1502 showed a negative association with production of aPPA and tended to reduce the number of total streptococci in saliva. In contrast, the DRB1* 1501 allele was significantly correlated with a high level of induction of the antibodies, and also tended to reduce lactobacilli and mutans streptococci. Further, peptide immunogenicity was confirmed in NOD-SCID mice grafted with human peripheral blood mononuclear cells. Our results indicate that the interplay between regulators such as age, DRB1 genotype, cytokines, and peptide immunogenicity may provide a potential means for developing a vaccine useful for the prevention of dental caries as well as their diagnosis. [ABSTRACT FROM AUTHOR]

Selective IgA deficiency is the most common form of primary immunodeficiency, the molecular basis of which is unknown. To investigate the cause of selective IgA deficiency, we examined what stage of B-cell differentiation was blocked. DNA and RNA were extracted from three Japanese patients with selective IgA deficiency and three with a partial IgA deficiency. In selective IgA deficiency patients, I α germline transcript expression levels decreased and α circle transcripts were not detected. Stimulation with PMA and TGF- β1 up-regulated I α germline and α circle transcripts. In some patients, IgA secretion was induced by stimulation with anti-CD40, IL-4 and IL-10. In partial IgA deficiency patients, I α germline, α circle transcripts and C α mature transcripts were detected in the absence of stimulation. Our findings suggest that the decreased expression level of I α germline transcripts before a class switch might be critical for the pathogenesis of some patients with selective IgA deficiency. However, in patients with a partial IgA deficiency, B-cell differentiation might be disturbed after a class switch. [ABSTRACT FROM AUTHOR]

We investigated, by Northern blotting, ELISA, and a chemotaxis assay, the expression of IL-8 mRNA, the production of IL-8 protein, and the biological activity of mononuclear cells (MNC), polymorphonuclear neutrophils (PMN) and plasma, respectively, from patients with Kawasaki disease (KD) who received intravenous immunoglobulin (IVIG). IL-8 mRNA expression by MNC and PMN, the level of IL-8 protein, and the neutrophil chemoattractant activity within plasma were all increased in the acute phase of KD, and were significantly elevated following IVIG therapy. The level of chemotactic activity of neutrophils, but not that of monocytes, in response to F-met-leu-phe was decreased in patients with KD after IVIG. The increased expression of IL-8 in PMN and MNC, the increased plasma level of IL-8 and the decreased level of neutrophil chemotactic activity of the patients who received IVIG therapy might inhibit the accumulation of neutrophils at the sites of inflammation, and may thus reduce the risk of aneurysm formation. [ABSTRACT FROM AUTHOR]

CD4+ cells from autoimmune-prone C57BL/6 lpr/lpr mice contain two subpopulations. B220- CD4+ and B220+CD4+ cells. Highly purified B220- CD4+ cells from C57BL/6 +/+ and lpr/lpr mice were examined by comparing functional characteristics and expression of cell surface antigens and T cell receptor (TcR)/CD3 complex. Both lpr B220+CD4+ and B220+CD4- CD8- cells, most of which were PgP-1 positive, expressed TcR/CD3 complex on the cell surface at lower level as compared with B220- CD4+ cells of age-matched normal mice. In addition, the B220- CD4+ cells were heterogeneous on the basis of surface expression of PgP-1 and CD3 antigens. Normal levels of TcR Cα-. Cβ- and Vβ8-specific mRNA were found in the B220-CD4+ cells and B220+CD4+ cells as compared with normal B220+CD4+ cells, while Vβ8-specific mRNA was preferentially expressed only by B220+CD4--CD8- cells. Either B220+CD4+ cells and B220+CD4- CD8- cells failed to respond to anti-CD3 monoclonal antibody (MoAb) as assessed by proliferative responses and production of interleukin-2 (IL-2). However, appreciable levels of reactivity to anti-CD3 MoAb were detected in the B220- CD4+ cells, although the responsiveness of this subset to such stimuli were reduced, compared with those of normal control. These results indicate that the B220- CD4+ cells in lpr mice are phenotypically and functionally distinct from normal B220-CD4+ cells. [ABSTRACT FROM AUTHOR]

Autoimmune-prone mice homozygous for the 1pr gene develop prominent lymphadenopathy composed mainly of Thy-1+ CD8- CD4- B220+ cells. Expression patterns of B220vsCD4 on lymph node cells from 1pr mice were analysed using two-colour flow microfluorometry. B220+CD4+ cells, which were hardly seen in lymph nodes of B6-+ / + mice, increased significantly in B6-lpr mice with ageing. Functional analysis of purified B220+ CD4+ cells from 1pr mice revealed that these cells scarcely responded to T cell mitogens with or without rIL-2. Furthermore, B220+ CD4+ cells were defective in IL-2 production when cultured with Con A. On the other hand, B220-CD4+ cells from B6-lpr mice showed an ability to respond to T cell mitogens similar to that of B220- CD4+ cells from B6-+/+ mice. These results indicate that an unusual T cell subset expressing both B220 and CD4 in 1pr mice is functionally defective, but the intrinsic ability of B220- CD4+ cells is almost intact as compared with the counterpart in normal mice. [ABSTRACT FROM AUTHOR]

An autosomal recessive gene, lpr, is responsible for lymphoproliferation and autoimmunity of lpr mice, in which background genes are also known to influence the development of autoimmune disease. To define the differences in abnormally proliferating T cells between C57BL/6-lpr/lpr and MRL/Mp-lpr/lpr mice, and to try and understand the influence of background in the differing expression of autoimmune disease in both strains, we analysed the sequences of T cell antigen receptor Vβ genes expressed in the cells from the enlarged lymph nodes of C57BL/6-lpr/lpr mice. Eleven β cDNAs out of the 38 Cβ-specific cDNAs contained sequences with open reading frames from the beginning of the variable region to the expected termination codons at the end of the constant regions. Notably, 36% of the functional β-chain mRNAs expressed Vβ8.3 genes, whereas Vβ8.3 and Vβ8.2 genes were not found. These results are consistent with a relatively lower frequency of the Vβ8.1 or Vβ8.2 expressing cells in the hypertrophic lymph nodes of C57BL/6-lpr/lpr mice, detected by KJI6-133 monoclonal antibody. Interestingly, other Vβ genes expressed in these mice were completely distinct from those in MRL/Mp-lpr/lpr mice as described by Singer et al (1986), The different distribution of Vβ genes expressed in C57BL/6-lpr/lpr from that in MRL/Mp-lpr/lpr mice might be related to the differences in the genetic background and the expression of lpr gene-associated autoimmunity. [ABSTRACT FROM AUTHOR]

Primary hyperparathyroidism associated with Albright's hereditary osteodystrophy was diagnosed in a 22 year old Japanese woman, the second such case to be reported. Albright's hereditary osteodystrophy (AHO) appears to be associated with a larger number of disorders than the well recognized pseudohypoparathyroidism. AHO and pseudo-pseudohypoparathyroidism are essentially identical. [ABSTRACT FROM PUBLISHER]

Objective It is not clear whether a 3-minute single– stage Adenosine tilt (ATP tilt) test is useful to diagnose patients with unexplained syncope. Isoproterenol (ISP) is often utilized to induce neurally mediated syncope (NMS). The aim of this study is to evaluate ATP tilt test and compare ATP tilt test with ISP tilt test. Methods and Results ATP tilt test and ISP tilt test were performed in 103 patients (66 men, mean age: 52.3±18.6 years) with unexplained syncope. After a drug-free tilt test of 30 minutes, subjects were administered 0.1mg/kg and/or 0.2mg/kg ATP in the upright position (tilt angle of 80°) and observed for up to 3 minutes. ISP was infused constantly and patients were observed for up to 15 minutes in the upright position. A positive ATP tilt was found in 29 (28.1%) of 103 patients and a positive ISP tilt in 5 (7.9%) of 63 patients. Both ATP tilt and ISP tilt were positive in only one (1.6%) of 63 patients. Patients with positive ATP tilt had characteristics statistically different from those with a positive ISP tilt. The diagnostic yield of ATP tilt was higher than that of ISP tilt. Conclusion An ATP tilt was effective in the diagnosis of NMS patients in whom ISP tilt was negative. [ABSTRACT FROM PUBLISHER]

Objectives Transthoracic epicardial ablation is an emerging catheter ablation strategy being used clinically at increasing frequency. This study evaluates the effects of epicardial RF ablation in the vicinity of the coronary arteries (CA) in the pigs. Methods The study consisted of 14 pigs. A total of 20 lesions were created in the vicinity of CA by cooling on and 23 lesions were created in the detached of CA. RF power was 30 W for 60 seconds by cooling-on. We studied the biophysical characteristics of RF ablation lesions and the damage of CA. Results In the vicinity of CA, effective RF procedure was 77 % (15/20) by cooling-on and 40 % (4/10) by cooling-off (P=0.06). In the vicinity of CA, lesions created with cooling-on and cooling-off RF ablation were 4.4±1.6mm and 1.8±0.3 mm in depth (P<0.01). In the detached of CA, lesions created with cooling-on and cooling-off RF ablation were 7.1±2.0 mm and 4.1±1.1mm in depth (P<0.05). In the vicinity of CA, lesions created with cooling-on and cooling-off RF ablation were 173±57 mm3 and 68±22 mm3 in volume (P<0.01). In the detached of CA, lesions created with cooling-on and cooling-off RF ablation was 422±118 mm3 and 125±23 mm3 in volume. There was no damage to major epicardial arteries when the catheter tip was 5 mm away from the arteries. Conclusions Cooling-on RF ablation is useful and safety if ablation sites are 5 mm away from CA in the vicinity of CA [ABSTRACT FROM PUBLISHER]

Background The recurrence rate after ablation of typical atrial flutter (AFL) is still higher than that of other supraventricular tachyarrhythmias. The purpose of the present study was to determine whether performing ablation using electro-anatomical mapping (CARTO) as guide is able to reduce the incidence of AFL recurrences during the follow-up. Methods Our study population comprises two groups of patients with AFL who underwent linear ablation of cavo-tricuspidal isthmus: 1) from 1996 to 2002, the bidirectional block at level of isthmus was confirmed by conventional method such as changes of propagation sequence on the Halo catheter and/or occurrence of double potential on ablation line (Conv group); 2) from 2002 to 2005, the bidirectional block was confirmed by CARTO system (CARTO group). Result We enrolled 150 consecutive patients (age 63.0±11.9 yrs). The follow-up period averaged 921 days. The recurrence rate of AFL of Conv group was 12.4% (n=14/113) during 1189 days, while the recurrence rate of CARTO group was 8.1% (n=3/37) during 140 days. In patients who experienced recurrences, the isthmus length was longer than in patients who did not (30 vs 24 mm). Conclusion The recurrence rate of AFL during the follow-up seems to be lower in patients in whom ablation was performed by means of CARTO system. An isthmus length greater than 30 mm is predictor of AFL recurrence. [ABSTRACT FROM PUBLISHER]