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CAP Home > CAP Reference Resources and Publications > CAP TODAY > CAP Today Archive 2002 > December 2002 Clinical Abstracts
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  Clinical Abstracts

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cap today

December 2002

Overexpression of the BAFF gene in Sjögren’s syndrome
Symptoms of dry mouth and dry eyes caused by malfunctioning exocrine glands, such as salivary and lacrimal glands, are characteristic of the chronic inflammatory disorder Sjögren’s syndrome. This autoimmune disease is characterized by large mononuclear cell infiltrates in exocrine glands, B-cell hyperreactivity, and various serum autoantibodies. Sjögren’s syndrome can develop alone or in association with other autoimmune diseases. Abnormal B-cell activity is a predominant feature of the condition, which is manifested by polyclonal B-cell activation and elevated secretion of autoantibodies, such as rheumatoid factors, anti-Ro (SS-A), anti-La (SS-B), and anti-α-fodrin autoantibodies. However, the role of B cells and autoantibodies in the pathogenesis of the condition remains unclear. The gene BAFF is a powerful modulator of B-cell biology expressed by monocytes/macrophages and dendritic cells. Expression of BAFF is B-cell specific. Overproduction of BAFF is known to be associated with the development of certain autoimmune diseases. Transgenic (Tg) mice with BAFF mutation have an elevated number of B cells in the periphery, secrete various autoantibodies, and develop a systemic lupus erythematosus-like condition. Excess BAFF-mediated survival signals might compromise the ability of autoreactive B cells to respond to censoring death signals; therefore, abnormal BAFF production may be a key event in autoimmunity. Serum BAFF levels are significantly higher in patients with SLE and rheumatoid arthritis than in healthy individuals. The authors described the findings in BAFF Tg mice. As the mice age, they develop a condition secondary to their lupus-like disease and some perceived similarities to Sjögren’s syndrome in humans. The animals show severe sialadenitis, decreased saliva production, and destruction of submaxillary glands. Sjögren’s syndrome in humans also correlates with elevated levels of circulating BAFF and dramatic upregulation of BAFF expression in inflamed salivary glands. Marginal zone B cells, one of the B cell subsets in the spleen of BAFF Tg mice, is a potential reservoir of autoreactive B cells. B cells with a marginal zone-like phenotypic pattern infiltrate the salivary glands of the BAFF Tg mice. The authors concluded that altered B cell differentiation and tolerance induced by excess BAFF may be a central component of the pathogenesis of Sjögren’s syndrome.

Groom J, Kalled SL, Cutler AH, et al. Association of BAFF/BLyS overexpression and altered B cell differentiation with Sjögren’s syndrome. J Clin Invest. 2002; 109: 59-68.

Reprints: Fabienne Mackay, The Garvan Institute of Medical Research, 384 Victoria St., Darlinghurst NSW 2010, Australia; f.mackay@garvan.unsw.edu.au

Storage artifact of clinical hematology specimens
Dyserythropoiesis occurs in a variety of conditions, including megaloblastic anemias, iron deficiency, thalassemias, congenital anemias, infections, toxic exposure, and preleukemic states. Observation of dyserythropoiesis using the French-American-British classification scheme is important in bone marrow interpretation. Nonetheless, dyserythropoiesis may be spurious and an artifact of storage in clinical specimens. One must be aware of this artifact to avoid erroneous diagnosis of myelodysplastic syndrome. The authors documented the occurrence and severity of dyserythropoiesis as an artifact of storage of clinical hematology specimens. They studied bone marrow aspirates collected in EDTA obtained from seven patients without myelodysplasia. These specimens were stored at room temperature (20 to 24°C) or refrigerated temperature (1 to 6°C) and examined for dyserythropoiesis at zero, one, two, and three days of storage. The initial specimens showed few abnormalities, but nuclear aberrations occurred in about one percent of the erythroid population. When stored at room temperature, dyserythropoietic changes increased significantly with each day of storage. Nuclear and cytoplasmic aberrations also occurred. The cytoplasmic changes were more extensive than the nuclear abnormalities. The mean percentage of erythroblasts with cytoplasmic vacuoles increased by day of storage as follows: day zero, ≈1.1 percent; day one, ≈22.2 percent; day two, ≈29.4 percent; day three, ≈35.6 percent. On day one, nuclear shaped changes increased to ≈6.21 percent, on day two to ≈11.36 percent, and on day three to ≈12.85 percent. The authors found that after one day of storage at room temperature, sufficient dysplastic changes occur to cause difficulty in diagnosing myelodysplastic syndrome. These changes appear to be inhibited significantly by refrigerated storage.

Wang LJ, Glasser L. Spurious dyserythropoiesis. Am J Clin Pathol. 2002;117:57-59.

Reprints: Dr. Lewis Glasser, Dept. of Pathology, Rhode Island Hospital, 593 Eddy St., Providence, RI 02903

Maternal serum assays for activin A and inhibin A in trisomy 18 pregnancies
The triple screen, involving assays for maternal serum α-fetoprotein, total human chorionic gonadotropin, and unconjugated oestriol, detects 60 percent of trisomy 18 pregnancies (false-positive rate, 0.2 to 0.7 percent). Levels of maternal serum pregnancy-associated plasma protein-A (PAPP-A) and free β-hCG are reduced in such pregnancies. And fetal nuchal translucency thickness algorithms have been developed that, along with the PAPP-A and free β-hCG measurements, identify 90 percent of cases (false-positive rate, one percent). Controversy surrounds the value of adding inhibin A measurement to second trimester screening protocols for trisomy 21. Limited data suggest that inhibin A cannot discriminate trisomy 18 aneuploidy in the second trimester of pregnancy. Likewise, activin has been found in various small studies to be a poor discriminator of trisomy 21. The authors of this study evaluated whether maternal serum levels of inhibin A and total activin A are altered in the first trimester of pregnancies involving trisomy 18. They studied 45 cases of trisomy 18 and 493 control pregnancies. The pregnancies were examined at 10 to 14 weeks of gestation. The authors measured serum inhibin A, total activin A, free β-hCG, and PAPP-A. The median values in the trisomy 18 pregnancies were 0.74 MoM for inhibin A, 1.23 MoM for activin A, 0.38 MoM for free β-hCG, and 0.16 MoM for PAPP-A. The levels of inhibin and activin deviated from normal only slightly in comparison with those for free β-hCG and PAPP-A. The authors concluded that adding these two assays to the screening for trisomy 18 is not likely to further improve the sensitivity of screening.

Spencer K, Liao AW, Ong CYT, et al. Maternal serum activin A and inhibin A in trisomy 18 pregnancies at 10-14 weeks. Prenat Diagn. 2001;21:571-574.

Reprints: Kevin Spencer, Endocrine Unit, Clinical Biochemistry Dept., Harold Wood Hospital, Gubbins Lane, Romford, Essex RM3 OBE, United Kingdom; kevinspencer1@cs.com

A prospective longitudinal study of gene polymorphisms in rheumatoid arthritis
HLA and non-HLA genes contribute to the pathogenesis of rheumatoid arthritis. The presence and dose effect of HLA-DRB1 alleles encoding the shared epitope affect the course and outcome of the disease. Estimates of the HLA component of overall genetic risk for RA are less than 50 percent. Identifying new susceptibility and severity genes for RA is an important challenge being addressed via genome-wide screening or candidate gene approach. Matrix metalloproteinases (MMP) have been implicated in connective tissue destruction and remodeling associated with various chronic disease conditions. The expression of these metalloproteinases is regulated at the transcription level by a variety of factors. Collagenase-1 (MMP-1) can degrade the interstitial collagens types I, II, and III. It is produced by synoviocytes and chondrocytes. The level of MMP-1 is elevated in the plasma and synovial fluid of patients with RA. The authors undertook a prospective longitudinal study to test the hypothesis of an association between the MMP-1 gene polymorphism and susceptibility to and severity of RA. They used the candidate gene approach in a cohort of 103 patients diagnosed early with RA and followed for four years. They also investigated the association between HLA-DRB1 gene polymorphism and the severity of RA. The authors used a radiographic damage score to quantify disease severity at baseline and after four years of followup in each patient. They used fluorescent-based polymerase chain reaction to analyze MMP-1 polymorphism genotyping. And they used PCR sequence-specific oligonucleotide probes for HLA-DRB1 genotyping. These assays were also performed on 133 healthy control subjects. The authors found that the MMP-1 allele and genotype frequencies did not differ between RA patients and controls. Radiographic damage or its progression over four years of followup did not differ across MMP-1 genotypes. The radiographic damage score and its progression over four years did, however, differ across HLA-DRB1 genotypes. The HLA-DRB1 shared epitope +/+ genotype was associated with the highest radiographic damage score, while the shared epitope -/- genotype was associated with the lowest. The authors concluded that their results do not support the hypothesis of an association between the MMP-1 gene promoter polymorphism and susceptibility to or severity of RA.

Constantin A, Lauwers-Cancès V, Navaux F, et al. Collagenase-1 (MMP-1) and HLA-DRB1 gene polymorphisms in rheumatoid arthritis: a prospective longitudinal study. J Rheum. 2002;29:15-20.

Reprints: A. Cantagrel, Service de Rhumatologie, CHU Rangueil, 31403 Toulouse Cedex 4, France; cantagre@cict.fr

Karyotype followup on maternal serum screening for Down syndrome
Several fetal chromosome anomalies, in addition to Down syndrome, are found in populations that are subjected to maternal serum screening, including trisomy 18, Turner Syndrome, and other aneuploidies. It is not clear which aneuploidies are concentrated in the group at increased risk for Down syndrome and which are present by chance. The authors used the databases of the South Australian State Neonatal Screening Programme, South Australian Maternal Serum Antenatal Screening (SAMSAS) Programme, and other sources to create a complete list of abnormal karyotypes present in the pregnancies screened at increased risk of fetal Down syndrome. There were 65,328 pregnancies screened by the SAMSAS Programme from 1991 through 1997. Of these, 5.25 percent were declared at increased risk of fetal Down syndrome, and 79.8 percent, including 16 with early fetal loss, had fetal or neonatal karyotypes determined. Overall, 129 instances of abnormal karyotype were found, including 84 Down syndrome, four trisomy 18, three trisomy 13, two triploidy, six female sex chromosome aneuploidy and five male sex chromosome aneuploidy, 19 inherited balanced rearrangements, four mosaic or de novo balanced abnormalities, and two unbalanced karyotypes. Of these, only the karyotype for Down syndrome occurred in the pregnancies at increased risk for fetal Down syndrome with a frequency greater than that expected for the general pregnant population.

Ryall RG, Callen D, Cocciolone R, et al. Karyotypes found in the population declared at risk of Down syndrome following maternal serum screening. Prenat Diagn. 2001;21:553-557.

Reprints: R.G. Ryall, Dept. of Chemical Pathology, Women’s & Children’s Hospital, North Adelaide, South Australia 5006, Australia; ryallr@wch.sa.gov.au

New modification of the Lancefield classification for group A streptococci
The Lancefield classification system for typing group A streptococci was developed in 1928. It is based on variable antigenic properties of protein M on the surface of the organisms. The authors reported on an exchange of strains between six international group A streptococci (GAS) reference laboratories that resulted in 22 additional sequence types designated emm103 to emm124 being added to the M protein gene sequence types. Continued expansion of the serology-based Lancefield classification scheme for GAS has become difficult during the past 20 years. Using a new sequence-based methodology that adheres to previously established strain criteria, while being predictive of known M protein serotypes, has permitted types emm94 to emm102 to be added to the Lancefield scheme. Continued expansion through the addition of emm103 to emm124 is now proposed. As with emm94 to emm102, each of these new emm types was represented by multiple independent isolates recovered from serious disease manifestations. Each of these was nontypeable with all M protein typing sera stocks available to international GAS reference laboratories. And each demonstrated antiphagocytic properties in vitro by multiplying in normal human blood.

Facklam RF, Martin DR, Lovgren M, et al. Extension of the Lancefield classification for group A streptococci by addition of 22 new M protein gene sequence types from clinical isolates: emm103 to emm124. Clin Infect Dis. 2002;34:28-38.

Reprints: Dr. Bernard Beall, Centers for Disease Control and Prevention, Mail Stop C02, 1600 Clifton Rd. NE, Atlanta, GA 30333; bbeall@cdc.gov

An assay for determining anti-acetylcholine receptor antibodies
The antigenic target in the autoimmune disorder myasthenia gravis is the nicotinic acetylcholine receptor (nAChR) of skeletal muscles. Diagnosing and monitoring this disease involves detecting and quantifying anti-acetylcholine receptor antibodies in serum. The most commonly used assay for this purpose is the immunoprecipitation assay using iodine125 labeled α-bungarotoxin (125I-α-BuTx). This is a valid, specific, and widely used assay; however, it is highly desirable to reduce the use of radioactivity in clinical laboratories. High-sensitivity detection of lanthanides uses time-resolved fluorescence methods. The authors developed a nonradioactive immunoassay using Eu3+-labeled α-cobratoxin and time-resolved fluorescence. They derivatized α-CTx with a diethylenetriaminepentaacetate moiety and formed a complex with Eu3+. This complex was purified by high-pressure liquid chromatography and the fractions tested for receptor binding. The Eu3+-labeled α-CTx competed with 125I-α-BuTx for binding to nicotinic acetylcholine receptors and saturated the binding sites of human acetylcholine receptors. The results of the immunoassays performed with Eu3+-labeled α-CTx were similar to those obtained with 125I-α-BuTx, with a slightly higher limit of detection (0.3 nmol/L versus 0.1 nmol/L for the isotopic assay). None of 34 negative sera tested gave a value greater than 0.3 nmol/L. Thirty-five positive myasthenic sera were compared using the two assays, and 32 tested positive with the Eu3+ assay. Linear regression analysis yielded the equation y=1.035x-0.013 nmol/L; Sy/x=0.172 nmol/L; and r2=0.977. The authors concluded that this new time-resolved fluorescence assay is comparable to the widely used isotopic assay.

Ricny J, Simkova L, Vincent A. Determination of anti-acetylcholine receptor antibodies in myasthenic patients by use of time-resolved fluorescence. Clin Chem. 2002;48:549-554.

Jan Ricny, Institute of Physiology, Academy of Sciences of Czech Republic. Videoska 1083, 142 20 Prague, Czech Republic; ricny@biomed.cas.cz

   
 

 

 

   
 
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