Studies on the types of immune responses to synthetic antigens in guinea-pigs

The immune response in guinea-pigs to linear and multichain copolymers of two, three or four different amino acids including tyrosine, glutamic acid, alanine and lysine, was studied.

Delayed-type response was favoured in the case of preparations of relatively low molecular weight, containing only tyrosine and glutamic acid in their potential antigenic specificity determinants. Delayed sensitivity to one of these preparations was passively transferred with lymphoid cells.

Antibody response resulted from the immunization of animals with preparations of relatively high molecular weight and with a more heterogeneous chemical composition. The antibody formation was intensified and accelerated, when p-azobenzenearsonate conjugates of the polypeptides were used as antigens.

Variations in the immunizing dose had little or no effect on the nature of the reactions. A response of exclusively delayed type could not be intensified by repeated immunizations. The presence of mycobacteria in the immunizing injection was essential for the elicitation of the response to multichain, but not to linear copolymers.

     

A paradigm for single nucleotide polymorphism analysis: the case of the acetylcholinesterase gene

Acetylcholinesterase (AChE) plays a crucial physiological role in termination of impulse transmission at cholinergic synapses through rapid hydrolysis of acetylcholine. It is a highly conserved molecule, and only a few naturally occurring genetic polymorphisms have been reported in the human gene. The goal of the present study was to make a systematic effort to identify natural single nucleotide polymorphisms (SNPs) in the human ACHE gene. To this end, the genomic coding sequences for acetylcholinesterase of 96 unrelated control individuals from three distinct ethnic groups were analyzed. A total of 13 ACHE SNPs were identified, 10 of which are newly described, and five that should produce amino acid substitutions [c.101G>A (p.Arg34Gln), c.169G>A (p.Gly57Arg), c.1031A>G (p.Glu344Gly), c.1057C>A (p.His353Asn), and c.1775C>G (p.Pro592Arg)]. Population frequencies of 11 of the 13 SNPs were established in four different populations: African Americans, Ashkenazi Jews, Sephardic Jews, and Israeli Arabs; 15 haplotypes and five ethnospecific alleles were identified. The low number of SNPs identified until now in the ACHE gene is ascribed to technical hurdles arising from the high GC content and the presence of numerous repeat sequences, and does not reflect its intrinsic heterozygosity. Among the SNPs resulting in an amino acid substitution, three are within the mature protein, mapping on its external surface: they are thus unlikely to affect its catalytic properties, yet could have antigenic consequences or affect putative protein-protein interactions. Furthermore, the newly identified SNPs open the door to a study of the possible association of AChE with deleterious phenotypes-such as adverse drug responses to AChE inhibitors employed in treatment of Alzheimer patients and hypersensitivity to pesticides.     

A new prenatal sonographic sign of epidermolysis bullosa

We report on the prenatal sonographic appearance of epidermolysis bullosa (EB). The third viable pregnancy of a consanguineous couple was found at 23 weeks to have dysplastic external ears and nose. The neonate was born at 33 weeks and was found to have junctional EB with pyloric atresia. On reviewing the 23‐week ultrasound images, skin denudation was evident. This is a report of visualization of skin denudation in EB. When EB is suspected prenatally, special attention should be given to the visualization of skin surfaces.     

Normal colon tissue and colon carcinoma show no difference in heparanase promoter methylation

Introduction

Heparanase, the sole heparan sulfate degrading enzyme, has a role in cellular invasion. Accordingly, a large number of studies have demonstrated an association between heparanase expression and tumor stage and patients' prognosis. In colon carcinoma, heparanase shows increased expression in tumor compared to normal tissue and its expression correlates with the presence of metastasis. One of the regulatory mechanisms of heparanase expression is de-methylation on its promoter. In the present study we evaluated the role of heparanase promoter methylation in colon carcinoma.

Material and methods

Analysis of heparanase promoter methylation was done on 32 samples of colon carcinoma as well as 30 samples of normal colonic mucosa. DNA was extracted from FFPE tissue and subjected to bisulfite conversion. The relative fraction of methylated and unmethylated DNA was evaluated using quantitative real-time PCR.

Results

The fraction of methylated DNA was 1 ± 3.4% in the colon carcinoma group, and 2.5 ± 3.3% in the normal colon group (P = 0.11). Only one case in the normal group and one case in the tumor group showed more than 10% methylation in the heparanase promoter.

Conclusion

We did not find any significant difference in heparanase promoter methylation between colon carcinoma and normal colonic mucosa, suggesting that heparanase overexpression in colon carcinoma is mediated by other mechanisms.     

Analysis of genetic polymorphisms in acetylcholinesterase as reflected in different populations

Acetylcholinesterase (AChE) plays a crucial physiological role in termination of impulse transmission at cholinergic synapses through rapid hydrolysis of acetylcholine. In addition, it was implicated in amyloid plaque formation, a hallmark of Alzheimer's disease (AD), and most of the drugs used in AD treatment are AChE inhibitors. Thus ACHE is an obvious candidate gene for pharmacogenetic study of AD treatment. However, AChE is a highly conserved molecule, and only a few naturally occurring genetic polymorphisms have been reported in the human gene. The goals of this study were to make a systematic effort to identify natural single nucleotide polymorphisms (SNPs) in the human ACHE gene, and to reveal their population specific architecture. To this end, the genomic coding sequences for AChE of 96 unrelated control individuals from three distinct ethnic groups, African Americans, Ashkenazi Jews and Israeli Arabs, were analyzed. Thirteen ACHE SNPs were identified, ten of which are newly described, and five of which should produce amino-acid substitutions (Arg34Gln, Gly57Arg, Glu344Gly, His353Asn and Pro592Arg). Population frequencies of 11 of the 13 SNPs were established in four different populations, African Americans, Ashkenazi Jews, Sephardic Jews and Israeli Arabs; 17 haplotypes and 5 ethno-specific alleles were identified, and a cladogram of ACHE haplotypes was constructed. Among the SNPs resulting in an amino-acid substitution, three are within the mature protein, mapping on its external surface; they are thus unlikely to affect its catalytic properties, yet could have antigenic consequences or affect putative protein-protein interactions. Furthermore, the newly identified SNPs open the door to a study of the possible association of AChE with deleterious phenotypes - such as adverse drug responses to AChE inhibitors employed in treatment of AD patients and hypersensitivity to pesticides.     

Plasma homocysteine level and functional outcome of patients with ischemic stroke

OBJECTIVE: To investigate the possible relationships between total plasma homocysteine level (tHcy) and functional outcome of stroke patients as evaluated by the FIM instrument. DESIGN: Retrospective chart analysis. SETTING: Inpatient stroke rehabilitation ward of a university-affiliated referral hospital. PARTICIPANTS: Consecutive patients (N=113) presenting with acute ischemic stroke. Patients were divided into 2 groups according to their tHcy levels (< or = 15 micromol/L, >15 micromol/L) and into 3 groups according to their FIM scores (low, < or =40; moderate, 41-80; high, >80). INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: The tHcy level was determined shortly after stroke onset by a high performance liquid chromatography method with fluorescence detection. Functional outcome was measured by the FIM instrument at admission and discharge. The tHcy level and FIM scores were obtained for all patients. Data outcomes were analyzed by t tests, 1-way analysis of variance, Mann-Whitney U, and Fisher exact tests, as well as by the 2 ordered polytomous logistic regression model. RESULTS: The 2 tHcy groups were similar in demographic, stroke, and comorbidity characteristics, differing only by higher frequency of hypertension in those with a tHcy greater than 15 micromol/L (51.7% vs 80.8%, respectively, P=.01). Compared with patients who had tHcy levels at 15 micromol/L or lower and were discharged from rehabilitation being in the highest FIM score group (>80), higher tHcy levels were not associated with a discharge FIM score of less than 40 (odds ratio [OR]=.77; 95% confidence interval [CI], 0.13-4.65; P=.77) or with a better functional outcome FIM score between 40 and 80 (OR=3.71; 95% CI, 0.73-18.99; P=.11). CONCLUSIONS: Our findings suggest that determination of tHcy level does not correlate with functional outcome in patients presenting for rehabilitation after acute ischemic stroke.     

Pregnancy outcome after in utero exposure to angiotensin converting enzyme inhibitors or angiotensin receptor blockers

ObjectiveTo examine first trimester safety of angiotensin-converting-enzyme-inhibitors (ACEIs) or angiotensin-receptor-blockers (ARBs).Study designProspective observational cohort regarding pregnancy ACEI/ARBs-exposure including contacts to two Teratology Information Services in Israel (1994–2007) and Italy (1990–2008), with two comparison groups: (1) exposed to other antihypertensives (OAH) (2) after non-teratogenic exposure (NTE) in similar time frames.Results252 ACEI/ARBs-exposed, 256 OAH-exposed and 495 NTE-exposed pregnancies were followed-up. The rate of major congenital anomalies was comparable between the groups (8/190, 4.2%, ACEI/ARB; 9/212, 4.2%, OAH; 18/471, 3.8% NTE; p = 0.954) among first trimester exposed pregnancies. The median gestational age at delivery was two weeks earlier, rate of preterm deliveries more than 2-fold higher, and median birth weight more than 200 g lower in the ACEI/ARB and OAH groups compared to the NTE group.ConclusionThe present study suggests that ACEI/ARBs are not major teratogens when used in the first trimester, and can reassure women with similar exposures.