Use of estimated breeding values in a selection index to breed Yorkshire pigs for high and low immune and innate resistance factors

Abstract

A random bred population of Yorkshire pigs (G_o) was characterized using fourteen various indicators of immune and innate resistance. Based on initial heritability estimates and correlations between these traits, two measures of antibody (serum IgG, and antibody response to HEWL), and cellular activity (blastogenic response to Con A and cutaneous DTH to BCG/PPD), and one indicator of innate monocyte function (uptake and killing of S. typhimurium) were chosen as breeding criteria to be used in a composite selection index. Based on these five traits a combined estimated breeding value (EBV) was calculated for each animal and pigs were assigned to High, Low or Control breeding groups. Approximately 120 first generation piglets (G₁) were then similarly evaluated. Based on G_o plus G₁ heritability estimates were 0.25, 0.23, 0.08, 0.08 and zero for secondary antibody response to HEWL, blastogenic response to Con A, cutaneous DTH to BCG/PPD, serum IgG, and monocyte function, respectively. Least squares means reflected these estimates in that there were significant (p ≤ 0.05) differences between High and Low line G₁ pigs in antibody, blastogenic, and DTH responses. However, there were no significant line differences in serum IgG or uptake and killing of S. typhimurium. Response to selection was determined both by differences in least squares means and differences in average EBV between the High and Low lines. After one generation of selection, lines were separated by 1.508 (least squares) and 1.205 (EBV) index points, or slightly more than half a standard (phenotypic) deviation.     

Sick Sinus Syndrome: Experience of a Cardiac Pacemaker Clinic

Out of a pacemaker clinic population of 182 patients 21 (11·5%) were found to have the sick sinus syndrome. Their ages ranged from 30 to 80 years and averaged 62. Ischaemic heart disease was more commonly an aetiological factor than in patients with chronic atrioventricular heart block. Other aetiologies were familial cardiomyopathy, postcardiac surgery, and dystrophia myotonica.Cardioversion before pacemaker insertion was a hazardous procedure. After insertion the occurrence of tachycardias and the need for drug therapy were reduced. One patient no longer required a pacemaker once atrial fibrillation became established.A high incidence of cerebral embolization was observed and the use of anticoagulant drugs therefore merits serious consideration.Failure of inhibition of demand-type pacemakers occurred in two patients. Two patients who exhibited competition with fixed-rate pacemakers died. Two patients were treated with electrodes surgically implanted on the right atrium. It is suggested that fixed-rate pacemakers may be contra-indicated and that sequential atrioventricular demand pacing is theoretically ideal.     

The RNA-binding protein Staufen1 is increased in DM1 skeletal muscle and promotes alternative pre-mRNA splicing

In myotonic dystrophy type 1 (DM1), dystrophia myotonica protein kinase messenger ribonucleic acids (RNAs; mRNAs) with expanded CUG repeats (CUG(exp)) aggregate in the nucleus and become toxic to cells by sequestering and/or misregulating RNA-binding proteins, resulting in aberrant alternative splicing. In this paper, we find that the RNA-binding protein Staufen1 is markedly and specifically increased in skeletal muscle from DM1 mouse models and patients. We show that Staufen1 interacts with mutant CUG(exp) mRNAs and promotes their nuclear export and translation. This effect is critically dependent on the third double-stranded RNA-binding domain of Staufen1 and shuttling of Staufen1 into the nucleus via its nuclear localization signal. Moreover, we uncover a new role of Staufen1 in splicing regulation. Overexpression of Staufen1 rescues alternative splicing of two key pre-mRNAs known to be aberrantly spliced in DM1, suggesting its increased expression represents an adaptive response to the pathology. Altogether, our results unravel a novel function for Staufen1 in splicing regulation and indicate that it may positively modulate the complex DM1 phenotype, thereby revealing its potential as a therapeutic target.     

Linkage disequilibrium detected between dystrophia myotonica and APOC2 locus in the Finnish population

Summary Three polymorphic loci APOC2, CKMM and p134C were used to haplotype 15 Finnish dystrophia myotonica (DM) families representing about one third of all DM patients in this isolated population. Compound APOC2 and CKMM haplotypes reveal linkage disequilibrium: 90% of DM chromosomes co-occur with the haplotypes that occur in 31% of normal chromosomes only. The same disequilibrium is present when only polymorphisms occurring at the APOC2 locus are used. Surprisingly, no statistically significant linkage disequilibrium was discovered at the CKMM locus alone. Of the meiotic events, 84% were informative when both APO2 and CKMM loci were used. When studied selectively, 60% of meiotic events were informative at the APOC2 locus, whereas CKMM alone resulted in 65% meiotic informativeness. The distal marker p134C was found to have an unfortunately low information content in our population.     

Effect of respiratory syncytial virus infection on the uptake of and immune response to other inhaled antigens

Groups of BALB/c mice were sham infected or inoculated intranasally (IN) with live RSV. From Day 4 to 8 after infection, the animals were exposed IN to ovalbumin (OVA) with or without alum adjuvant. At different intervals, levels of OVA concentration in serum, IgG-anti-OVA antibody activity in serum, and IgA-anti-OVA antibody activity in bronchial washings were determined, employing the ELISA technique. IgE-anti-OVA antibody titers in serum and bronchial washings were assessed by PCA. OVA concentrations in serum were significantly higher in RSV-infected animals compared to uninfected controls. The use of alum adjuvant also increased OVA uptake in uninfected animals but to a lesser extent than RSV infection. RSV-infected animals developed significantly higher OVA-specific antibody titers of IgG isotype in serum and IgA isotype in bronchial washings than the uninfected controls, while alum enhanced the immune response less markedly but still significantly in uninfected mice. An IgE antibody response to OVA in serum was demonstrable in 50% of RSV-infected mice immunized IN with OVA and alum, while all uninfected animals and RSV-infected animals immunized with OVA alone (without adjuvant) failed to develop a detectable IgE response. These findings suggest that infections with viral agents such as RSV may function as adjuvants for other antigens inhaled during acute respiratory infection. These observations may explain the alterations in the immune response to other antigens in patients with acute viral-induced bronchopulmonary diseases.     

Malaria eradication: the economic,financial and institutional challenge

Background

Plasmodium falciparum infection causes cerebral malaria (CM) in a subset of patients with anti-malarial treatment protecting only about 70% to 80% of patients. Why a subset of malaria patients develops CM complications, including neurological sequelae or death, is still not well understood. It is believed that host immune factors may modulate CM outcomes and there is substantial evidence that cellular immune factors, such as cytokines, play an important role in this process. In this study, the potential relationship between the antibody responses to the merozoite surface protein (MSP)-1 complex (which consists of four fragments namely: MSP-1₈₃, MSP-1₃₀, MSP-1₃₈ and MSP-1₄₂), MSP-6₃₆ and MSP-7₂₂ and CM was investigated.

Methods

Peripheral blood antibody responses to recombinant antigens of the two major allelic forms of MSP-1 complex, MSP-6₃₆ and MSP-7₂₂ were compared between healthy subjects, mild malaria patients (MM) and CM patients residing in a malaria endemic region of central India. Total IgG and IgG subclass antibody responses were determined using ELISA method.

Results

The prevalence and levels of IgG and its subclasses in the plasma varied for each antigen. In general, the prevalence of total IgG, IgG1 and IgG3 was higher in the MM patients and lower in CM patients compared to healthy controls. Significantly lower levels of total IgG antibodies to the MSP-1_f38, IgG1 levels to MSP-1_d83, MSP-1₁₉ and MSP-6₃₆ and IgG3 levels to MSP-1_f42 and MSP-7₂₂ were observed in CM patients as compared to MM patients.

Conclusion

These results suggest that there may be some dysregulation in the generation of antibody responses to some MSP antigens in CM patients and it is worth investigating further whether perturbations of antibody responses in CM patients contribute to pathogenesis.     

α2-Macroglobulin-proteinase complexes in cystic fibrosis serum Analysis by monoclonal antibodies and receptor-mediated endocytosis by normal human fibroblasts

α₂-Macroglobulin complexed to proteinases activated during clotting of cystic fibrosis and control sera was quantitated with the complex-specific monoclonal antibody F₂B₂. Similar amounts of α₂-macroglobulin complexes (between 40 and 90 μg/ml) were generated in cystic fibrosis and control sera. Endocystosis of the complexes by normal fibroblasts was compared to the amount of complexes detected by the F₂B₂-radioimmunoassay. Normal uptake was observed with 13 out of 14 cystic fibrosis sera. One cystic fibrosis serum showed strongly reduced endocytosis of the complexes. Complexes isolated from this serum on immobilized F₂B₂ failed to inhibit binding of purified α₂-macroglobulin-trypsin to its receptor, demonstrating deficient receptor-binding of these complexes. The low complexes could not be distinguised from complexes isolated from control or other cystic fibrosis sera by isoelectric focusing, rate electrophoresis or SDS-polyacrylamide gel electrophoresis.     

Viral Antibody Levels and Clinical Status in Acute Exacerbations of Chronic Bronchitis: a Controlled Prospective Study

A controlled prospective study was made of a group of patients with chronic bronchitis, in which serum antibodies against a group of viruses and Mycoplasma pneumoniae were estimated at regular intervals. Sixteen significant rises in antibody titre were shown, of which eight were associated with clinical acute exacerbations of bronchitis. In individual patients no correlation was found between the number of acute exacerbations or the aetiological agent and persistent change in ventilatory function as expressed by the F.E.V._0.75.This study was compared with the results of a previous parallel investigation of the same patients done to study the significance of rhinovirus infections. In the present investigation 12% of the acute exacerbations were associated with the 11 agents tested compared with 14% associated with rhinoviruses in the earlier work.     

Selenium and its relationship with selenoprotein P and glutathione peroxidase in children and adolescents with Hashimoto’s thyroiditis and hypothyroidism

The essential trace element selenium (Se) is required for thyroid hormone synthesis and metabolism. Selenoproteins contain selenocysteine and are responsible for biological functions of selenium. Glutathione peroxidase (GPx) is one of the major selenoproteins which protects the thyroid cells from oxidative damage. Selenoprotein P (SePP) is considered as the plasma selenium transporter to tissues. The aim of this study was to evaluate serum Se and SePP levels, and GPx activity in erythrocytes of children and adolescents with treated Hashimoto’s thyroiditis, hypothyroidism, and normal subjects.Blood samples were collected from 32 patients with Hashimoto’s thyroiditis, 20 with hypothyroidism, and 25 matched normal subjects. All the patients were under treatment with levothyroxine and at the time of analysis all of the thyroid function tests were normal. GPx enzyme activity was measured by spectrophotometry at 340 nm. Serum selenium levels were measured by high-resolution continuum source graphite furnace atomic absorption. SePP, TPOAb (anti-thyroid peroxidase antibody), and TgAb (anti-thyroglobulin antibody) were determined by ELISA kits. T₄, T₃, T₃ uptake and TSH were also measured.Neither GPx activity nor SePP levels were significantly different in patients with Hashimoto’s thyroiditis or hypothyroidism compared to normal subjects. Although GPx and SePP were both lower in patients with hypothyroidism compared to those with Hashimoto’s thyroiditis and normal subjects but the difference was not significant. Serum Se levels also did not differ significantly in patients and normal subjects. We did not find any correlation between GPx or SePP with TPOAb or TgAb but SePP was significantly correlated with Se.Results show that in patients with Hashimoto’s thyroiditis or hypothyroidism who have been under treatment with levothyroxine and have normal thyroid function tests, the GPx, SePP and Se levels are not significantly different.     

Expanded CUG repeats trigger aberrant splicing of ClC-1 chloride channel pre-mRNA and hyperexcitability of skeletal muscle in myotonic dystrophy

In myotonic dystrophy (dystrophia myotonica, DM), expression of RNAs that contain expanded CUG or CCUG repeats is associated with degeneration and repetitive action potentials (myotonia) in skeletal muscle. Using skeletal muscle from a transgenic mouse model of DM, we show that expression of expanded CUG repeats reduces the transmembrane chloride conductance to levels well below those expected to cause myotonia. The expanded CUG repeats trigger aberrant splicing of pre-mRNA for ClC-1, the main chloride channel in muscle, resulting in loss of ClC-1 protein from the surface membrane. We also have identified a similar defect in ClC-1 splicing and expression in two types of human DM. We propose that a transdominant effect of mutant RNA on RNA processing leads to chloride channelopathy and membrane hyperexcitability in DM.     

Modifications to toxic CUG RNAs induce structural stability,rescue mis-splicing in a myotonic dystrophy cell model and reduce toxicity in a myotonic dystrophy zebrafish model

CUG repeat expansions in the 3′ UTR of dystrophia myotonica protein kinase (DMPK) cause myotonic dystrophy type 1 (DM1). As RNA, these repeats elicit toxicity by sequestering splicing proteins, such as MBNL1, into protein–RNA aggregates. Structural studies demonstrate that CUG repeats can form A-form helices, suggesting that repeat secondary structure could be important in pathogenicity. To evaluate this hypothesis, we utilized structure-stabilizing RNA modifications pseudouridine (Ψ) and 2′-O-methylation to determine if stabilization of CUG helical conformations affected toxicity. CUG repeats modified with Ψ or 2′-O-methyl groups exhibited enhanced structural stability and reduced affinity for MBNL1. Molecular dynamics and X-ray crystallography suggest a potential water-bridging mechanism for Ψ-mediated CUG repeat stabilization. Ψ modification of CUG repeats rescued mis-splicing in a DM1 cell model and prevented CUG repeat toxicity in zebrafish embryos. This study indicates that the structure of toxic RNAs has a significant role in controlling the onset of neuromuscular diseases.     

Production of antibodies specific to human chorionic gonadotropin in mice immunized against its chemical analogs

Mice immunized against DS₅-hCG-Β and DS₆-hCG-Β, chemical analogs of Β-subunit of human choriogonadotropin (hCG-Β) in which 5 and 6 disulphide bonds respectively were reduced and alkylated, were found to produce antibodies specific to hCG without significant crossreactivity with human lutropin (hLH) as tested in a radioimmunoassay. In contrast, mice immunized against the native hCG-Β subunit produced hLH crossreacting antibodies. While the anti-DS5, DS6-hCG-Β serum was capable of selectively blocking the binding of [¹²⁵I]-hCG to rat testicular LH/hCG receptors, it failed to inhibit the binding of [₁₂₅I]-hLH to the same receptors. The radioimmunoassay for hCG using the mouse anti-DS₅, DS₆-hCG-Β serum was not as sensitive as that employing rabbit anti-DS₅, DS₆-hCG-Β serum. The minimal detection limit was 5 ng/ml for the mouse antibody as compared to 1 ng/ml for the rabbit antibody. Supported by U.S. Public Health Service Grant HD 08766 to OPB. An erratum to this article is available at .     

Inhibition of Nitrate Transport by Anti-Nitrate Reductase IgG Fragments and the Identification of Plasma Membrane Associated Nitrate Reductase in Roots of Barley Seedlings

Membrane associated nitrate reductase (NR) was detected in plasma membrane (PM) fractions isolated by aqueous two-phase partitioning from barley (Hordeum vulgare L. var CM 72) roots. The PM associated NR was not removed by washing vesicles with 500 millimolar NaCl and 1 millimolar EDTA and represented up to 4% of the total root NR activity. PM associated NR was stimulated up to 20-fold by Triton X-100 whereas soluble NR was only increased 1.7-fold. The latency was a function of the solubilization of NR from the membrane. NR, solubilized from the PM fraction by Triton X-100 was inactivated by antiserum to Chlorella sorokiniana NR. Anti-NR immunoglobulin G fragments purified from the anti-NR serum inhibited NO₃⁻ uptake by more than 90% but had no effect on NO₂⁻ uptake. The inhibitory effect was only partially reversible; uptake recovered to 50% of the control after thorough rinsing of roots. Preimmune serum immunoglobulin G fragments inhibited NO₃⁻ uptake 36% but the effect was completely reversible by rinsing. Intact NR antiserum had no effect on NO₃⁻ uptake. The results present the possibility that NO₃⁻ uptake and NO₃⁻ reduction in the PM of barley roots may be related.     

Adjuvant and immunogenic properties of bacterial lipopolysaccharide in IgE and IgG1 antibody formation in mice

The ability of Gram-negative lipopolysaccharide (LPS) to function as an adjuvant and as an antigen in IgE and IgG₁ immune responses in mice was investigated. LPS failed to induce LPS-specific IgE or IgG₁ under a variety of experimental conditions. Both isolated LPS and whole heat-killed bacteria were capable of enhancing IgE and IgG₁ antibody formation to a protein antigen, egg albumin (EA). The LPS-induced anti-EA, IgE, and IgG₁ antibody titers exhibited a cycling phenomenon with time. In the presence of LPS, IgE, and IgG₁ antibodies specific for EA did not occur in athymic nude (BALB/c-nu/nu) mice, demonstrating the inability of LPS to substitute for the stringent requirement for T cells in homocytotropic antibody formation.     

The PPARγ agonist rosiglitazone promotes the induction of brite adipocytes,increasing β-adrenoceptor-mediated mitochondrial function and glucose uptake

Recruitment and activation of brite (or beige) adipocytes has been advocated as a potential avenue for manipulating whole-body energy expenditure. Despite numerous studies illustrating the differences in gene and protein markers between brown, brite and white adipocytes, there is very little information on the adrenergic regulation and function of these brite adipocytes. We have compared the functional (cyclic AMP accumulation, oxygen consumption rates, mitochondrial function, glucose uptake, extracellular acidification rates, calcium influx) profiles of mouse adipocytes cultured from three contrasting depots, namely interscapular brown adipose tissue, and inguinal or epididymal white adipose tissues, following chronic treatment with the peroxisome proliferator-activated receptor γ (PPARγ) agonist rosiglitazone. Prototypical brown adipocytes readily express β₃-adrenoceptors, and β₃-adrenoceptor stimulation increases cyclic AMP accumulation, oxygen consumption rates, mitochondrial function, glucose uptake, and extracellular acidification rates. Treatment of brown adipocytes with rosiglitazone increases uncoupling protein 1 (UCP1) levels, and increases β₃-adrenoceptor mitochondrial function but does not affect glucose uptake responses. In contrast, inguinal white adipocytes only express UCP1 and β₃-adrenoceptors following rosiglitazone treatment, which results in an increase in all β₃-adrenoceptor-mediated functions. The effect of rosiglitazone in epididymal white adipocytes, was much lower compared to inguinal white adipocytes. Rosiglitazone also increased α₁-adrenoceptor mediated increases in calcium influx and glucose uptake (but not mitochondrial function) in inguinal and epididymal white adipocytes. In conclusion, the PPARγ agonist rosiglitazone promotes the induction and function of brite adipocytes cultured from inguinal and epididymal white adipose depots.     

慢性阻塞性肺疾病急性加重期患者血清SAA水平与肺功能、炎性因子的相关性及其诊断价值分析

目的:探讨慢性阻塞性肺疾病急性加重期(AECOPD)患者血清淀粉样蛋白A(SAA)水平与肺功能及炎性因子的相关性,并分析其诊断价值。方法:选取2013年6月-2018年6月中国人民解放军第970医院收治的204例慢性阻塞性肺疾病(COPD)患者作为研究对象,其中COPD稳定期患者132例作为COPD稳定组,AECOPD患者72例作为AECOPD组。另选取同期于中国人民解放军第970医院进行健康体检的50例健康体检者作为健康组。比较三组受试者血清SAA水平、白细胞介素-8(IL-8)、白细胞介素-6(IL-6)、C反应蛋白(CRP)、降钙素原(PCT)水平及肺功能,采用Pearson相关性分析AECOPD患者血清SAA水平与肺功能及炎性因子的相关性,并分析SAA对AECOPD的诊断价值。结果:AECOPD组患者血清SAA、PCT、CRP、IL-6、IL-8水平较COPD稳定组及健康组升高(P0.05),COPD稳定组患者血清SAA、IL-6、IL-8、PCT、CRP水平均高于健康组,差异有统计学意义(P0.05);AECOPD组患者第一秒用力呼气容积(FEV1)、用力肺活量(FVC)、第一秒用力呼吸容积占用力肺活量的百分比(FEV1/FVC%)、第一秒用力呼气容积占预计值百分比(FEV1%)低于COPD稳定组及健康组,差异有统计学意义(P0.05),COPD稳定组患者FEV1、FVC、FEV1/FVC%、FEV1%低于健康组,差异有统计学意义(P0.05)。Pearson相关性分析显示,AECOPD患者血清SAA水平与IL-8、IL-6、CRP、PCT呈正相关,与FEV1%、FEV1/FVC%、FEV1、FVC呈负相关(P0.05)。受试者工作特征(ROC)曲线结果显示,SAA对AECOPD诊断的敏感度为80.85%,特异度为80.07%,曲线下面积为0.832。结论:AECOPD患者血清SAA水平明显升高,其与患者肺功能及炎症因子存在相关性,具有较高的诊断价值,可用于AECOPD患者病情的评估。     

Evaluation of test immunisation in the assessment of antibody deficiency syndromes.

Antibody responses after immunisation with pneumococcal polysaccharide did not correlate with the severity and frequency of infections in 22 patients with severe hypogammaglobulinaemia, when these were measured by a Farr radioimmunoassay. Five "healthy" patients with severe hypogammaglobulinaemia not only failed to make antipneumococcal polysaccharide antibody, when measured by radioimmunoassay, but also had very low or unrecordable antibody responses to Escherichia coli and failed to produce antibody when immunised with tetanus toxoid. Some of these subjects, however, did make small amounts of IgM antipneumococcal polysaccharide antibody when this was measured by an enzyme linked immunosorbent assay, while others retained some ability to produce IgM or IgA or both in their saliva. These findings show that the measurement of serum antibody responses after immunisation, with the possible exception of IgM antibodies to polysaccharides, is unlikely to be helpful in assessing the requirement for gammaglobulin replacement therapy in patients with hypogammaglobulinaemia.     

Hybridoma growth in a new generation hollow fibre bioreactor: antibody productivity and consistency

This paper analyses the performance of MAbMax^TM/Tricentric^TM, a new generation hollow fibre bioreactor, for hybridoma growth and antibody productivity, the down stream processing of monoclonal antibody harvests throughout the run and the further control of antibody quality consistency. Handling and process parameters were optimised using a mouse hybridoma, IgG1_K secretor, and then confirmed with several other hybridomas. Cells were kept at optimal viability during an unusually long period of time and a continuously high production of antibodies was detected over several months. Foetal bovine serum concentration was reduced to 1\% and the effects of weaning of cells from serum were monitored in terms of cell metabolism and antibody productivity. Antibody harvests collected at regular intervals throughout the run (2 to 12 weeks) were purified using affinity chromatography on a recombinant protein A/G matrix and then analysed in terms of antigen binding properties, isoelectric forms and oligosaccharide structures, in order 1) to control antibody quality consistency as a function of time and serum concentration and 2) to compare antibody characteristics as a function of culture conditions, in vitro bioreactor cultivation versus in vivo mouse ascite cultivation. This revised version was published online in July 2006 with corrections to the Cover Date.