College of American Pathologists
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  Clinical Abstracts





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March 2002

  • Immunoglobulin subclass patterns in recurrent miscarriages
  • Effect of air pollution on plasma fibrinogen levels
  • Type II heparin-induced thrombocytopenia
  • Leukocyte leukotrienes as biomarkers for survival in critical patients
  • Serum lipid and lipoprotein levels in type 1 diabetic mothers and fetuses
  • Comparison of multiplex PCR and viral isolation for identifying HSV
  • Assessment of vitamin status in neonates
  • D-dimer as a marker of acute bowel ischemia

    Immunoglobulin subclass patterns in recurrent miscarriages
    The authors examined IgG subclass patterns in normal pregnancy and recurrent miscarriage. Four patient groups were studied. Group one was composed of 10 healthy nonpregnant women. Group two was composed of 10 women in their first trimester of pregnancy whose pregnancies went to term. Group three was composed of eight women in their first trimester who had a history of at least three previous miscarriages and whose pregnancies during the study went to term. Group four was composed of 10 women in their first trimester who had at least three previous miscarriages and whose pregnancies again failed in the first trimester. Serum IgG subclasses were measured by an enzyme-linked immunosorbent assay method. Women with a normal pregnancy (group two) had significant increases in total IgG and IgG 1, 2, and 3 in early pregnancy compared with women who were not pregnant. Group three did not differ significantly from group two. Women in group four showed significantly reduced levels of total IgG and IgG 1, 2, 3, and 4 when compared with women in group two. IgG 1 levels were significantly higher in patients in group three than in group four. This study demonstrated that normal pregnancy is associated with significant changes in IgG subclasses and that pregnant women who subsequently miscarried in the first trimester showed different IgG subclass patterns. IgG 1 levels were significantly increased in ongoing pregnancies, while miscarriage was associated with significantly reduced IgG 1 levels. The authors noted that changes leading to miscarriage occur at an early stage of pregnancy, and they hypothesized that various cytokines may be responsible for the IgG subclass patterns observed.

    Wilson R, Maclean MA, Jenkins C, et al. Abnormal immunoglobulin subclass patterns in women with a history of recurrent miscarriage. Fertil Steril. 2001;76:915-917.

    Correspondence: Dr. Rhoda Wilson, Dept. of Medicine, Glasgow Royal Infirmary, 10 Alexandra Parade, Glasgow G31 2ER, Scotland, United Kingdom;

    Effect of air pollution on plasma fibrinogen levels
    Cardiovascular morbidity and mortality have been associated with air pollution levels in several cities worldwide. The reason for this association is not known. The authors examined the hypothesis that the link may be the effect of air pollution on plasma fibrinogen levels. The authors measured plasma fibrinogen levels daily in 4,982 male and 2,223 female office workers in London between September 1991 and May 1993 and compared these findings with data on the concentration of various air pollutants during the day of the blood sample and the preceding three days. The data were adjusted for weather and other confounding factors. Major changes in NO2 and CO levels (from the 210th to 90th percentiles) were associated with a 1.5 percent increase in plasma fibrinogen. These relationships were more pronounced in the warmer season (April to September). Similar associations of plasma fibrinogen with black smoke and particulate matter occurred only in the warm season. SO2 and ozone showed no association. The authors suggested that ultrafine particles in urban air pollution may promote inflammation of pulmonary epithelium with associated acute phase responses.

    Pekkanen J, Brunner EJ, Anderson HR, et al. Daily concentrations of air pollution and plasma fibrinogen in London. Occup Environ Med. 2000;57:818-822.

    Reprints: Dr. Juha Pekkanen, Unit of Environmental Epidemiology, National Public Health Institute, P.O. Box 95, FIN-70701 Kuopio, Finland;

    Type II heparin-induced thrombocytopenia
    A major side effect of heparin therapy is thrombocytopenia. The moderate form of thrombocytopenia appears and spontaneously reverses over a few days with continued treatment. The more severe type II heparin-induced thrombocytopenia occurs five to 15 days after onset of heparin therapy. The latter has a more severe reduction of platelets, can result in thromboembolism, and normalizes only after withdrawal of heparin. The diagnosis of HIT can be difficult to establish clinically, so laboratory confirmation with biologic or antigen assays is necessary. The authors investigated whether laboratory confirmation can be improved after antigen clearance by combining the two types of assays. They collected blood from 14 HIT patients over five to six weeks and performed fluorescence-linked immunofiltration as the antigenic assay and carbon-14 serotonin release as the biologic assay. Eleven of the 14 patients were positive on day one post-heparin (sensitivity, 80 percent for each assay) using both assays. The three patients who were initially negative seroconverted during the next seven days, increasing the sensitivity to 100 percent at day seven. Both assays became negative on all patients within five to six weeks.

    Harenberg J, Wang LC, Hoffmann U, et al. Laboratory diagnosis of heparin-induced thrombocytopenia type II after clearance of platelet factor 4/heparin complex. J Lab Clin Med. 2001;137:408-413.

    Reprints: Dr. J. Harenberg, Dept. of Medicine IV, University Hospital, Theodor-Kutzer-Ufer, 68167 Mannheim, Germany

    Leukocyte leukotrienes as biomarkers for survival in critical patients
    ICU patients with severe systemic inflammation and sepsis show very high rates of morbidity and mortality. A large number of proinflammatory mediator molecules have been implicated in the pathophysiologic mechanism of systemic inflammatory response syndrome (SIRS). Evidence indicates that cysteinyl-leukotriene (leukotriene C4; LTC4), which is derived from arachidonic acid by the action of 5-lipoxygenase in leukocytes, may be one of these molecules. The authors studied 14 patients admitted to a hospital ICU in Germany who were found to have SIRS, sepsis, or sepsis syndrome. The study involved nine male and five female patients aged 42 to 82 years, as well as five healthy volunteers—three men and two women aged 34 to 38 years. The capacity of Ca-ionophore-stimulated leukocytes to synthesize LTC4 was measured by high-pressure liquid chromatography and radioimmunoassay. All patients initially synthesized less LTC4 than controls. Those who did not survive had continued to show low LTC4, whereas the levels returned to normal during the same observation period for those who survived.

    Morlion BJ, Torwesten E, Kuhn KS, et al. Cysteinyl-leukotriene generation as a biomarker for survival in the critically ill. Crit Care Med. 2000;28:3655-3658.

    Reprints: Dr. Peter Fürst, Institute for Biological Chemistry & Nutrition, University of Hohenheim, Garbenstr. 30, 70593 Stuttgart, Germany;

    Serum lipid and lipoprotein levels in type 1 diabetic mothers and fetuses
    Because type 1 diabetes mellitus is associated with alterations in lipid levels and changes in serum lipoproteins, the authors hypothesized that diabetes during pregnancy may further alter lipoprotein metabolism. Previous work has been inconsistent in demonstrating a relationship between lipid and lipoprotein alterations and type 1 diabetic pregnancies. This may have been because the degree of metabolic control of the diabetes, as determined by HbA1c levels, was not carefully studied. The authors investigated whether lipoprotein metabolism was altered by type 1 diabetes mellitus in pregnant women and whether there was a relationship between maternal and fetal lipoprotein metabolism. Mothers with poorly controlled or well-controlled type 1 diabetes, as indicated by HbA1c concentrations, had serum lipid, apolipoprotein, and lipoprotein lipid concentrations determined. Lipid measurements were also performed for their macrosomic (body weight =4,650±90 g) or appropriate-for-gestational-age newborns (body weight=3,616±68 g). These levels were compared with those of healthy mothers and their control newborns (body weight=3,290±45 g). The mothers and newborns with well-controlled diabetes showed serum lipid, apolipoprotein, and lipoprotein lipid concentrations comparable to those of the control mothers and newborns. The mothers with poorly controlled diabetes had higher serum triglyceride and apoprotein B-100 levels and lower apo A-1 and HDL3 cholesterol and lower phospholipid levels relative to the controls. In the macrosomic newborns, all serum lipid, apolipoprotein, and lipoprotein lipid levels were higher than for the control newborns. The authors concluded that degree of metabolic control is an important factor in whether type 1 diabetes affects maternal and fetal lipid levels.

    Merzouk H, Madani S, Korso N, et al. Maternal and fetal serum lipid and lipoprotein concentrations and compositions in type 1 diabetic pregnancy: relationship with maternal glycemic control. J Lab Clin Med. 2000;136:441-448.

    Reprints: Dr. J. Belleville, Faculté des Sciences Mirande, BP 400, 21011, Dijon Cedex, France

    Comparison of multiplex PCR and viral isolation for identifying HSV
    Herpes simplex virus type 2 and, less commonly, type 1, are the major causes of genital herpes. This infection typically manifests itself as crops of painful, disfiguring lesions that chronically recur. HSV infection recently has been implicated as an important cofactor in the acquisition and transmission of HIV. HSV is also a well-known etiologic agent in cases of encephalitis, aseptic meningitis, atypical pneumonias, and a variety of other infections. Laboratory diagnosis of genital herpes traditionally has relied on HSV cell culture as the gold standard. These cell culture techniques are quite sensitive and have proven to be particularly reliable in early stage genital infections. Direct antigen enzyme immunoassays and immunofluorescent techniques have also been used in identifying and serotyping HSV infections. These techniques have significantly reduced turnaround time with respect to cell culture. The turnaround time with these methods is approximately six hours versus two to seven days. The tradeoff, however, is that EIA is much less sensitive than traditional cell culture as a diagnostic tool. Both techniques have proven to be less sensitive as herpes lesions progress to crusting, healing, and reactivation. The authors investigated a polymerase chain-reaction assay for HSV as a diagnostic method for routine use in the clinical laboratory. They also considered cost and compared it with that for viral isolation. The authors studied patient sample swabs from 100 individuals that were inoculated onto a MRC-5 cell culture tube. The positive results on these cultures were confirmed by direct fluorescent antibody technique, and serotyping was performed using HSV-1 and -2 type-specific sera when requested. The PCR techniques used an extraction step of the same initial swab specimen followed by PCR amplification using a multiplex assay for HSV-1 and HSV-2 DNA. HSV-positive results were found in 32 of 100 samples via viral cultures and in 36 of 100 samples via PCR. The PCR positive results yielded 44 percent of patients infected with HSV-1 and 20 percent infected with HSV-2. The turnaround time for viral culture averaged 108 hours for positive results and 154 hours for negative results, whereas the PCR turnaround time averaged 24 to 48 hours. The cost to the laboratory of using viral culture for this diagnosis was $3.22 for a negative result and $6.49 for a positive result. Serotyping, when requested, added an additional $10.88 to each culture-positive test. The laboratory costs for PCR were higher, at $8.20 per sample, but the test was also reimbursed at a higher level. The authors concluded that it is feasible to use multiplexed PCR for the diagnosis of HSV-1 and -2 from patient swabs in routine clinical laboratory settings. The assay offers increased sensitivity and typing and improved turnaround time compared with viral culture techniques. Although the cost is high, the assay may be favorable in some situations.

    Marshall DS, Linfert DR, Draghi A, et al. Identification of herpes simplex virus genital infections: comparison of a multiplex PCR assay and traditional viral isolation techniques. Mod Pathol. 2001;14:152-156.

    Reprints: G.J. Tsongalis, Dept. of Pathology & Lab Medicine, Hartford Hospital, 80 Seymour St., Hartford, CT 06102;

    Assessment of vitamin status in neonates
    Fetuses, neonates, and pregnant women are in a state of rapid cell turnover that requires a high rate of DNA synthesis. In turn, this rapid growth and accompanying DNA synthesis are associated with a greater need for the water-soluble vitamins B-12, folate, and B-6. Early detection of vitamin deficiencies in these groups is important because the neurological changes that take place as a result of B-12 deficiency in infants may be irreversible. Functional, intracellular deficiency of these vitamins may occur in adults with minimal biochemical changes. Neuropsychiatric disorders resulting from B-12 deficiency may not be associated with typical clinical signs of anemia and macrocytosis. Serum vitamin concentrations thus have relatively poor sensitivity and specificity for detecting individuals with subtle changes suggestive of vitamin deficiency. Metabolites associated with such deficiencies, however, such as methylmalonic acid, homocysteine, cystathionine, and 2-methylcitric acid, have been found to be more sensitive estimates of functional and intracellular concentration deficiencies of these vitamins. Homocysteine, in particular, has been widely regarded as a reliable indicator for this purpose in adults, but there is little or no data indicating whether serum homocysteine is useful for monitoring vitamins in neonates and young infants. The authors studied total serum homocysteine levels in neonates and explored the relationship of these levels to serum vitamin concentrations, cysteine concentrations, and nutritional factors. They performed measurements of total homocysteine, cysteine, folate, vitamin B-12, and vitamin B-6 on 123 healthy, breast-fed neonates. The influence of the source of nutrition (formula versus human milk) was investigated in 60 infants. The mean total homocysteine concentration was 7.8±3.1 µmol/L. The total homocysteine concentration was linearly associated with the log rhythm of vitamin B-12 concentration (r= -0.64; P<0.001) and was also related to red blood cell folate concentration
    (r= -0.33; P<0.001) and cysteine concentrations (r= 0.36; P<0.001). The authors found more neonates with probable tissue deficiencies of vitamin B-12 and folate on the basis of total homocysteine measurements than had been expected from the analysis of serum vitamin concentrations alone (15.4 percent prevalence versus 9.7 percent predicted). Higher serum total homocysteine and cysteine concentrations with corresponding lower vitamin B-12 concentrations were found in breast-fed infants. The authors concluded that total homocysteine can be used as a functional indicator of vitamin B-12 and folate status in neonates. They also examined and recommended the use of the ratio of cysteine to vitamin B-12 as an additional index of impaired intracellular homocysteine metabolism.

    Minet JC, Bissé E, Aebischer CP, et al. Assessment of vitamin B-12, folate and vitamin B-6 status and relation to sulfur amino acid metabolism in neonates. Am J Clin Nutr. 2000;72:751-757.

    Reprints: E. Bissé, Dept. of Clinical Chemistry, University Hospital Freiburg, Hugstetterstrasse 55, D-79106 Freiburg, Germany; bissé

    D-dimer as a marker of acute bowel ischemia
    Prognosis of acute bowel ischemia secondary to occlusion of the superior mesenteric artery is a critical condition with a high mortality rate. Clinical presentation, white blood cell count, and serum amylase measurements are not helpful in the diagnosis. The decision point, clinically, relates to doing immediate laparotomy or angiography with subsequent thrombolysis. Laboratory signs, such as plasma lactate concentration and leucocytosis, have low diagnostic specificities. Metabolic acidosis is a late complication and, therefore, is not helpful. Contrast-enhanced computed tomography and plain radiography also are not specific. Duplex sonography can be useful, but the method is operator-dependent. The authors investigated the fibrinolytic marker D-dimer in the context of acute bowel ischemia. They analyzed 14 patients suspected of having acute bowel ischemia. Six of these patients had thromboembolic occlusion of the superior mesenteric artery, and all had significantly higher D-dimer levels than those without occlusion (P<0.05). Four patients with adhesions causing strangulated bowel, and one with ruptured aortic aneurysm, also had elevated D-dimer values. The authors concluded that more extensive prospective followup study is needed.

    Acosta S, Nilsson TK, Björck M. Preliminary study of D-dimer as a possible marker of acute bowel ischaemia. Br J Surg. 2001;88:385-388.

    Reprints: Dr. S. Acosta, Dept. of Surgery, Blekinge Hospital Karlskrona, SE 371 85 Karlskrona, Sweden;