Anti-calreticulin antibodies and calreticulin in sera of patients diagnosed with dilated or hypertrophic cardiomyopathy

Distinct cellular level of the Ca²⁺-binding chaperone calreticulin (CRT) is essential for correct embryonal cardiac development and postnatal function. However, CRT is also a potential autoantigen eliciting formation of antibodies (Ab), whose role is not yet clarified. Immunization with CRT leads to cardiac injury, while overexpression of CRT in cardiomyocytes induces dilated cardiomyopathy (DCM) in animals. Hence, we analysed levels of anti-CRT Ab and calreticulin in the sera of patients with idiopatic DCM and hypertrophic cardiomyopathy (HCM). ELISA and immunoblot using human recombinant CRT and Pepscan with synthetic, overlapping decapeptides of CRT were used to detect anti-CRT Ab. Serum CRT concentration was tested by ELISA. Significantly increased levels of anti-CRT Ab of isotypes IgA (p?p?p?相似文献   

Evidence for functional,inhibitory, histamine H3 receptors in rat carotid body Type I cells

The Type I cells are the sensory elements of the carotid bodies and play a critical role in defining the ventilatory response to hypoxia and hypercapnia. Type I cells release multiple neurotransmitters during a chemosensory stimulus resulting in increased firing of the carotid sinus nerve and modification of the breathing pattern. While much is known about the actions of individual neurotransmitters in this system, very little is known about how multiple neurotransmitters may integrate to shape the output of the carotid body. Recent data has indicated that the neurotransmitter histamine does not excite isolated Type I cells despite being released during hypoxia and its receptors being present on the Type I cells. Here the hypothesis that histamine might modulate an excitatory neurotransmitter such as acetylcholine was tested. Using calcium imaging techniques it was found that histamine attenuated calcium signaling events initiated by the muscarinic acetylcholine receptor agonist acetyl-β-methylcholine via an H3 receptor mediated mechanism. In summary, these results suggest that when acetylcholine and histamine are co-released from Type I cells in response to chemostimuli, histamine may attenuate or modulate the excitatory presynaptic actions of acetylcholine.     

Monitoring of autoantibodies against CYP4Z1 in patients with colon,ovarian, or prostate cancer

We have previously monitored the detection of autoantibodies (aAbs) directed against CYP4Z1 in the sera of breast and lung cancer patients. In the present study, the occurence of anti-CYP4Z1 aAbs in patients suffering from colon (n = 100), ovarian (n = 72), or prostate (n = 85) cancer was examined. Determination of aAbs was done using our previously established ELISA method. On average, the levels of anti-CYP4Z1 aAbs detected in sera from all cancer patients were not significantly higher than controls. No correlations were found with respect to gender or tumor stage. However, a subgroup of colon cancer patients with increased anti-CYP4Z1 aAb titers exhibited positive fecal occult blood test (FOBT) results and higher levels of both carbohydrate antigen 19–9 (CA19-9) and carcinoembryonic antigen (CEA). These results do not suggest that anti-CYP4Z1 aAbs have value as an independent biomarker for the detection of either colon, ovarian, or prostate cancer. However, they might be useful in combination with other biomarkers for the identification of a subset of colon cancers. Investigations involving a more powered sample size of this subgroup are needed to support this notion.     

Anti-mitochondrial antibodies in patients with dilated cardiomyopathy (anti-M7) are directed against flavoenzymes with covalently bound FAD

Anti-mitochondrial antibodies (anti-M7) in sera from patients with dilated cardiomyopathy and myocarditis recognize, besides mitochondrial antigens, bacterial sarcosine dehydrogenase. The common target antigen was identified as the covalently bound FAD of mitochondrial and bacterial flavoenzymes. Thus, anti-M7-positive serum reacted on Western blots exclusively with covalently flavinylated enzymes. The antigenic specificity of anti-M7 sera was reproduced by an antiserum raised in rabbits with 6-hydroxy- D -nicotine oxidase. The heart mitochondrial membrane antigen recognized by anti-M7 serum was identified as the flavoprotein subunit of succinate dehydrogenase, the antigens in rat liver mitochondrial matrix as the flavoenzymes dimethylglycine dehydrogenase and sarcosine dehydrogenase. Anti-M7 serum contained a specific anti-flavoenzyme antibody fraction. Nanomolar concentrations of FAD and riboflavin inhibited the immune reaction on Western blots and in ELISA, and incubation with FAD-agarose depleted the anti-M7 activity of the serum. N-terminally deleted dimethylglycine dehydrogenase proteins were only immunoprecipitated by anti-M7 sera when the FAD was covalently incorporated. An affinity constant (K_D) of 10^?8 M was established for the anti-flavoenzyme antibodies by competitive ELISA. Of patients with cardiomyopathy and myocarditis, 36% and 25%, respectively, were anti-flavoenzyme-positive by Western blot and ELISA, but only two of 15 patients with other heart diseases and none of 50 healthy controls.     

Chromosomal localization of the 5-HT1F receptor gene: No evidence for involvement in response to sumatriptan in migraine patients

The 5-HT_1F receptor, which is present in both human vascular and neuronal tissue, may mediate the therapeutic effect and/or side-effects of sumatriptan. We investigated the chromosomal localization of the 5-HT_1F receptor gene and the relation between eventually existing polymorphisms and the clinical response to sumatriptan in migraine patients. The 5-HT_1F receptor gene was localized using a monochromosomal mapping panel, followed by a radiation-reduced hybrid mapping and fluorescent in situ hybridization. The results of these techniques show that the 5-HT_1F receptor gene is localized at 3p12. We investigated the presence of polymorphisms by single strand conformation polymorphism analysis in 14 migraine patients who consistently responded well to sumatriptan, 12 patients who consistently experienced recurrence of the headache after initial relief, 12 patients with no response to sumatriptan, and in 13 patients who consistently experienced chest symptoms after use of sumatriptan. No polymorphisms were detected in any of the patients. We therefore conclude that genetic diversity of the 5-HT_1F receptor gene is most probably not responsible for the variable clinical response to sumatriptan. Am. J. Med. Genet. 77:415–420, 1998. © 1998 Wiley-Liss, Inc.     

Myocardial fibrosis in patients with symptomatic obstructive hypertrophic cardiomyopathy: correlation with echocardiographic measurements, sarcomeric genotypes, and pro-left ventricular hypertrophy polymorphisms involving the renin-angiotensin-aldosterone system

IntroductionHypertrophic cardiomyopathy (HCM) is a heterogeneous disorder of the cardiac sarcomere, resulting in myocyte hypertrophy and disarray, interstitial fibrosis, and cardiac dysfunction. Our aim was to determine whether the amount of fibrosis in HCM correlates with echocardiographic measures of diastolic dysfunction, presence of HCM-susceptibility mutations, or polymorphisms in the renin-angiotensin-aldosterone system (RAAS).MethodsSurgical specimens from patients with obstructive HCM undergoing septal myectomy at the Mayo Clinic (2001–2004) were examined and compared with autopsy-derived tissues from age- and sex-matched normal controls. Digital image analysis was used to quantitate the fibrosis in representative microscopic sections. Genotyping was performed for myofilament-HCM using polymerase chain reaction, high-performance liquid chromatography, and direct DNA sequencing. RAAS polymorphism status was similarly established.ResultsThe study included 59 HCM cases and 44 controls. Patients with HCM exhibited more fibrosis (mean 17%, range 3–45%) than controls (mean 8%, range 3–17%) (P<.0001). A significant relationship existed between amount of fibrosis and maximum wall thickness (P=.02), left ventricular ejection fraction (P=.02), and peak early/late diastolic mitral annulus velocity (E/A ratio) (P=.002). Although there was no association between amount of fibrosis and myofilament-HCM genotype status or polymorphisms in the RAAS cascade, there was a trend toward more fibrosis in patients with ≥1 C-encoding allele in CYP11B2-encoded aldosterone synthase.ConclusionsPatients with HCM undergoing septal myectomy had significantly more myocardial interstitial fibrosis than controls. The amount of fibrosis in HCM patients correlated with degree of septal hypertrophy and left ventricular systolic and diastolic function. Notably, neither mutations in cardiac myofilament proteins or polymorphisms in RAAS exhibited strong associations with severity of myocardial fibrosis.     

Anti-cytokine autoantibodies in autoimmunity: preponderance of neutralizing autoantibodies against interferon-alpha,interferon-omega and interleukin-12 in patients with thymoma and/or myasthenia gravis

We have screened for spontaneous anticytokine autoantibodies in patients with infections, neoplasms and autoimmune diseases, because of their increasingly reported co-occurrence. We tested for both binding and neutralizing autoantibodies to a range of human cytokines, including interleukin-1alpha (IL-1alpha), IL-1beta, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12, IL-18, interferon-alpha2 (IFN-alpha2), IFN-omega, IFN-beta, IFN-gamma, tumour necrosis factor alpha (TNF-alpha), transforming growth factor beta-1 (TGF-beta1) and granulocyte-macrophage colony stimulating factor (GM-CSF), in plasmas or sera. With two notable exceptions described below, we found only occasional, mostly low-titre, non-neutralizing antibodies, mainly to GM-CSF; also to IL-10 in pemphigoid. Strikingly, however, high-titre, mainly IgG, autoantibodies to IFN-alpha2, IFN-omega and IL-12 were common at diagnosis in patients with late-onset myasthenia gravis (LOMG+), thymoma (T) but no MG (TMG-) and especially with both thymoma and MG together (TMG+). The antibodies recognized other closely related type I IFN-alpha subtypes, but rarely the distantly related type I IFN-beta, and never (detectably) the unrelated type II IFN-gamma. Antibodies to IL-12 showed a similar distribution to those against IFN-alpha2, although prevalences were slightly lower; correlations between individual titres against each were so modest that they appear to be entirely different specificities. Neither showed any obvious correlations with clinical parameters including thymoma histology and HLA type, but they did increase sharply if the tumours recurred. These antibodies neutralized their respective cytokine in bioassays in vitro; although they persisted for years severe infections were surprisingly uncommon, despite the immunosuppressive therapy also used in most cases. These findings must hold valuable clues to autoimmunizing mechanisms in paraneoplastic autoimmunity.     

Structural and functional complexity of the humoral response against the Trypanosoma cruzi ribosomal P2 beta protein in patients with chronic Chagas' heart disease

High levels of antibodies against the C-terminus of the Trypanosoma cruzi TcP2 beta ribosomal protein, defined by the peptide EEEDDDMGFGLFD, named R13, have been measured in sera from patients with chronic Chagas' Heart Disease (cChHD). These antibodies also recognize an epitope on the second extracellular loop of the beta 1-adrenergic receptor, inducing a functional response on cardiomyocytes. The aim of this study was to gain novel insights into the structural basis of this cross-reactivity as well as to evaluate the origin of anti-M2- cholinergic receptor antibodies, which are also commonly found in cChHD patients. To address these questions we immunopurified anti-R13 antibodies and studied the structural requirements of epitope recognition. Results showed that the immunopurified antibodies recognized a conformation of R13 in which the third Glu residue was essential for binding, explaining their low affinity for the mammalian homologue (peptide H13: EESDDDMGFGLFD). Alanine mutation scanning showed individual variations in epitope recognition in each of the studied patients. The importance of a negatively charged residue at position 3 for the recognition of anti-R13 antibodies was further confirmed by competition experiments using a Ser3-phosphorylated H13 analogue, which had 10 times more affinity for the anti-R13 antibody than the native H13 peptide. Moreover, anti-R13 antibodies stimulated either the beta 1-adrenergic or the M2-cholinergic receptor, in strict agreement with the functional properties of the IgG fractions from which they derived, demonstrating that the same parasite antigen may generate antibody specificities with different functional properties. This may be a clue to explain the high variability of electrophysiological disturbances found in cChHD.     

Ursodeoxycholic acid treatment lowers the serum level of antibodies against pyruvate dehydrogenase and influences their inhibitory capacity for the enzyme complex in patients with primary biliary cirrhosis

A two-year randomized, double-blind, placebo-controlled clinical trial used paired serum samples from 122 patients with primary biliary cirrhosis to compare the effect of ursodeoxycholic acid and colchicine on their immune parameters. IgG antibodies to pyruvate dehydrogenase, the major autoantigen in primary biliary cirrhosis, were determined by enzyme-linked immunosorbent assay and immunoblot; enzyme inhibition assay against pyruvate dehydrogenase was used to test the changes of the functional reactivity of the serum autoantibodies. Treatment with ursodeoxycholic acid decreased both the level of IgG antibodies to pyruvate dehydrogenase (P<0.01) and the inhibitory titer of the sera for pyruvate dehydrogenase (P<0.01). Treatment with colchicine or placebo showed no statistically significant changes in either the antibody levels or the inhibitory titers. Ursodeoxycholic acid thus alters the immune parameters of patients with primary biliary cirrhosis. The mechanism of these changes needs further investigation.Abbreviations AMA Anti-mitochondrial antibodies - PBC Primary biliary cirrhosis - OD Optical density - PDH Pyruvate dehydrogenase - UDCA Ursodeoxycholic acid     

高血压病患者心脏指数、外周阻力及外周血液淋巴细胞β受体的检测

本实验应用放射配体结合法和超声心动图法分别测定了稳定型高血压病患者的外周血液淋巴细胞β受体密度和亲合力以及心脏指数和外周总阻力,并与正常人组进行了比较。结果显示高血压患者外周血液淋巴细胞β受体密度比正常人组值低(238±107对318±130 fmol/10~7细胞,P<0.05);二组之受体与配体复合物的平衡解离常数KD分别为2.62±1.25,3.31±1.14nm,二组间无显著差异(P>3.05)。高血压患者的心脏指数尚无显著性降低(3.92±1.06对4.54±1.09 L/min/M~2,P>0.05)而外周总阻力则明显增高(2811±711对1798±484 dynes sec.cm~(-3).M~2,P<0.01)。鉴于外周血液淋巴细胞β受体密度与心肌β受体密度呈正相关且受内源性儿茶酚胺的调节,本结果提示在稳定性高血压阶段也必然存在着心肌β受体密度的下降。我们认为这可能是高血压时尽管心脏所接受的肾上腺激素能刺激增加,但心排出量却不相应地增加甚至有下降趋势的原因之一。     

No evidence of mutations in the follicle‐stimulating hormone receptor gene in Mexican women with 46,XX pure gonadal dysgenesis

In the ovary FSH is necessary for normal follicular development, binding to its receptor (FSHR) that pertains to the superfamily of G‐protein coupled receptors. In the FSHR gene, which consists of 10 exons, an homozygous mutation was reported in six Finnish families with gonadal dysgenesis; whereas two isolated French patients exhibited compound heterozygous mutations. Several groups, however, have searched for FSHR mutations, although in most cases the gene has been studied partially, not finding any genetic abnormalities in German, English, North American or Brazilian women. We performed direct sequencing of all 10 exons of the FSHR gene in seven sporadic patients and two sisters with 46,XX pure gonadal dysgenesis, to investigate the cause of their disorder. No heterozygous or homozygous mutant alleles were present in any of the patients. Although the number of patients evaluated was small, considering all the other previous reports, it seems that except in the Finnish population, the proportion of women with mutations in the encoding region of this gene is very low. Other possibilities for the presence of 46,XX gonadal dysgenesis, such as defects in the regulatory regions of the FSHR gene promoter, in the untranslated regions of exons 1 and 10, and within introns, or the existence of other genes likely to be important for normal ovarian function on the X chromosome or on autosomes, should be considered. In contrast with other studies, we did not find polymorphisms of the FSHR gene, indicating that apparently in Mexicans this gene is not highly polymorphic. © 2001 Wiley‐Liss, Inc.     

No evidence for chromosomal mosaicism in multiple tissues of 10 patients with 45 XO Turner syndrome

Why the frequency of spontaneous abortions among monosomy X conceptuses is 98 % while the postnatal course of Turner syndrome is relatively benign has not been understood. One explanation could be that mosaicism for a euploid cell line confers viability and that those 2 % of 45, XO zygotes surviving in utero have some degree of mosaicism. We thus reasoned that if the non-mosaic 45, XO karyotype is lethal, a thorough study of living Turner syndrome patients might reveal a much higher frequency of mosaicism than the 30–40 % reported. Ten adult women with a 45 , XO leukocyte karyotype were investigated, looking at five tissue types from all three germ layers: buccal mucosa and hair from ectoderm, urinary epithelium from endoderm and ectoderm, and lymphocytes and skin fibroblasts from mesoderm. We were unable to confirm mosaicism in these patients, although in 2 out of 10 there was the suggestion of a small percentage of euploid cells in skin and blood karyotypes.     

阻塞性睡眠呼吸暂停综合征合并慢性阻塞性肺病 患者血清中β1与M2受体自身抗体的变化

目的: 检测阻塞性睡眠呼吸暂停综合征(OSAS)合并慢性阻塞性肺病(COPD)即重叠综合征(overlap syndrome,OS)患者血清中β₁－肾上腺素能受体与M₂－胆碱能受体自身抗体的变化。方法: 应用酶联免疫吸附法(ELISA)检测OS患者26例、OSAS患者32例、COPD患者30例及正常人28例血清中抗β₁与M₂受体的自身抗体。结果: β₁与M₂受体的自身抗体阳性率及滴度,OS组(92.2%, 57.7%及1:98, 1:67)明显高于OSAS组(71.9%, 40.6%及1:83, 1:30 )和COPD组(70.0%,36.7%及1:79, 1:28)(P＜0.05),OSAS和COPD组显著高于正常对照组(25.0%,14.3%及1:20, 1:20)(P＜0.01)。结论: OS,OSAS与COPD患者血清β₁与M₂受体自身抗体明显高于正常人,且以OS患者为著。     

Associations between genes for killer immunoglobulin-like receptors and their ligands in patients with epithelial ovarian cancer

Killer immunoglobulin-like receptors (KIRs) regulate function of NK cells and subsets of T cells. HLA class I molecules are ligands for inhibitory KIRs while specificity of activating KIRs is mainly unknown. Both KIR and HLA genotypes are highly polymorphic.     

No evidence of family history as a risk factor for herpes zoster in patients with post‐herpetic neuralgia

Little is known about reactivation of latent varicella zoster virus as herpes zoster in individuals with no underlying immunosuppression. Risk factors include age, sex, ethnicity, exogenous boosting of immunity from varicella contacts, underlying cell‐mediated immune disorders, mechanical trauma, psychological stress, and immunotoxin exposure. An association between herpes zoster and family history of zoster has been proposed. A case‐control study involving patients affected by post‐herpetic neuralgia, which usually follows more severe acute herpes zoster, was performed. The patients with post‐herpetic neuralgia were enrolled at the Pain Clinic of the Policlinico Tor Vergata in Rome, Italy, within 1 year from the onset of acute zoster. The controls matched for sex and age were chosen among healthy subjects without a history of herpes zoster presenting at the Internal Medicine Outpatient Clinic for hypertension in the same time period. All the participants in the study gave informed consent and were interviewed by medically trained and blinded investigators using a questionnaire. Similar proportions of the patients and the controls reported a family history of herpes zoster irrespective of the degree of relationship, i.e., 17.4% and 18.2%, respectively, by analyzing only the first‐degree relatives [RR 1.03 (CI 95%: 0.78–1.37)], and 28.4% and 29.6%, respectively, by analyzing the total number of relatives [RR 1.03 (CI 95%: 0.81–1.31)]. Further and larger prospective cohort studies are needed to ascertain whether a family history of herpes zoster is really an independent predictor of zoster in different geographical settings. J. Med. Virol. 82:1007–1011, 2010. © 2010 Wiley‐Liss, Inc.     

A triple antibody assay for the quantitation of plasma IgG subclass antibodies to acetylcholine receptors in patients with myasthenia gravis

Monoclonal anti-human IgG subclass antibodies have been used in an immunoprecipitation assay for the determination of anti-acetylcholine receptor IgG subclasses in plasma from patients with myasthenia gravis. Solubilized acetylcholine receptors labelled with 125I-alpha-bungarotoxin were incubated with patient plasma. Monoclonal mouse antibodies to human IgG subclasses 1-4 were added to the incubation and finally precipitated with anti-mouse IgG antibody. A maximal IgG subclass precipitation of 62-76% was determined with 125I-labelled myeloma IgG subclasses 1-4 added to normal human plasma. The anti-IgG subclass antibodies were added in excess which ensured that the precipitation of IgG2, IgG3 or IgG4 were unchanged, and that of IgG1 was only reduced by 17%, when the plasma IgG concentration was increased by a factor of two. The anti-IgG subclass antibodies were highly specific for their complementary subclasses. Determination of the IgG subclass of the anti-acetylcholine receptor antibodies from 8 patients with myasthenia gravis showed that IgG1 and IgG3 antibodies are predominant. This may support the hypothesis that complement mediated lysis of the neuromuscular end-plate plays a pathogenetic role in myasthenia gravis.     

Differential expression of the nuclear receptors farnesoid X receptor (FXR) and pregnane X receptor (PXR) for grading dysplasia in patients with Barrett's oesophagus

van de Winkel A, van Zoest K P M, van Dekken H, Moons L M G, Kuipers E J & van der Laan L J W
(2011) Histopathology 58 , 246–253
Differential expression of the nuclear receptors farnesoid X receptor (FXR) and pregnane X receptor (PXR) for grading dysplasia in patients with Barrett’s oesophagus Aims: To investigate expression of nuclear receptors farnesoid X receptor (FXR) and pregnane X receptor (PXR) as a diagnostic tool to improve grading of dysplasia in Barrett’s oesophagus patients. Methods and results: Immunostaining was analysed on a total of 192 biopsy samples of 22 Barrett’s patients with no dysplasia (ND), 17 with low‐grade dysplasia (LGD), 20 high‐grade dysplasia (HGD) and 24 with adenocarcinoma (AC). Nuclear FXR expression was observed in 15 of 22 (68%) ND cases versus none of 19 HGD; 3 of 17 (18%); LGD; 5 of 60 (8%) patients with AC (P < 0.001). FXR expression was highly specific for non‐dysplastic tissue. Nuclear PXR was expressed in 16 of 20 (80%) HGD cases versus two of 16 (13%) LGD cases (PPV 89%). Upon examining adjacent tissue taken from HGD and AC patients, PXR expression was high in samples of all tissue types. Conclusions: Nuclear receptors are expressed differentially during neoplastic progression, with FXR positivity being useful to distinguish ND from dysplasia and AC. PXR nuclear expression is able to separate HGD from LGD and ND. The combination of FXR and PXR also appears to have diagnostic and possibly prognostic value, but future prospective studies are required to investigate their predictive power for neoplastic progression in Barret’s oesophagus.