Saira Salahuddin

Angiogenic Factors and the Risk of Adverse Outcomes in Women with Suspected Preeclampsia

Background— An imbalance in circulating angiogenic factors plays a central role in the pathogenesis of preeclampsia. Methods and Results— We prospectively studied 616 women who were evaluated for suspected preeclampsia. We measured plasma levels of antiangiogenic soluble fms-like tyrosine kinase 1 (sFlt1) and proangiogenic placental growth factor (PlGF) at presentation and examined for an association between the sFlt1/PlGF ratio and subsequent adverse maternal and perinatal outcomes within 2 weeks. The median sFlt1/PlGF ratio at presentation was elevated in participants who experienced any adverse outcome compared with those who did not (47.0 [25th–75th percentile, 15.5–112.2] versus 10.8 [25th–75th percentile, 4.1–28.6]; P<0.0001). Among those presenting at <34 weeks (n=167), the results were more striking (226.6 [25th–75th percentile, 50.4–547.3] versus 4.5 [25th–75th percentile, 2.0–13.5]; P<0.0001), and the risk was markedly elevated when the highest sFlt1/PlGF ratio tertile was compared with the lowest (odds ratio, 47.8; 95% confidence interval, 14.6–156.6). Among participants presenting at <34 weeks, the addition of sFlt1/PlGF ratio to hypertension and proteinuria significantly improved the prediction for subsequent adverse outcomes (area under the curve, 0.93 for hypertension, proteinuria, and sFlt1/PlGF versus 0.84 for hypertension and proteinuria alone; P=0.001). Delivery occurred within 2 weeks of presentation in 86.0% of women with an sFlt1/PlGF ratio ≥85 compared with 15.8% of women with an sFlt1/PlGF ratio <85 (hazard ratio, 15.2; 95% confidence interval, 8.0–28.7). Conclusions— In women with suspected preeclampsia presenting at <34 weeks, circulating sFlt1/PlGF ratio predicts adverse outcomes occurring within 2 weeks. The accuracy of this test is substantially better than that of current approaches and may be useful in risk stratification and management. Additional studies are warranted to validate these findings.

Genomic polymorphism, growth properties, and immunologic variations in human herpesvirus-6 isolates

Fifteen human herpesvirus-6 (HHV-6) isolates from normal donors and patients with AIDS, systemic lupus erythematosis, chronic fatigue syndrome, collagen-vascular disease, leukopenia, bone marrow transplants, Exanthem subitum (roseola), and atypical polyclonal lymphoproliferation were studied for their tropism to fresh human cord blood mononuclear cells, growth in continuous T cell lines, reactivity to monoclonal antibodies, and by restriction enzyme banding patterns. All isolates replicated efficiently in human cord blood mononuclear cells, but mitogen stimulation of the cells prior to infection was required. The ability to infect continuous T-cell lines varied with the isolates. Isolates similar to GS prototype infected HSB2 and Sup T1 cells and did not infect Molt-3 cells, whereas isolates similar to Z-29 infected Molt-3 cells but not HSB2 and Sup T1 cells. Some of the monoclonal antibodies directed against the HHV-6 (GS) isolate showed reactivity with all isolates tested, but others only reacted with HHV-6 isolates similar to the GS isolate and not with those similar to Z-29 isolate. Restriction enzyme analysis using EcoRI, BamHI, and HindIII revealed that HHV-6 isolates from roseola, bone marrow transplant, leukopenia, and an HIV-1-positive AIDS patient from Zaire (Z-29) were closely related but distinct from GS type HHV-6 isolates. Based on the above findings, we propose that, like herpes simplex virus types 1 and 2, the 15 HHV-6 isolates analyzed can be divided into group A (GS type) and group B (Z-29 type).

Human B-lymphotropic virus (human herpesvirus-6).

Human B-lymphotropic virus (HBLV), also known as human herpesvirus-6 (HHV-6) was first isolated in 1986 from AIDS patients and patients with other lymphoproliferative disorders. HBLV is distinct from known human herpesviruses, biologically, immunologically and by molecular analysis. HBLV can infect and replicate in fresh and established lines of hemopoietic cells and cells of neural origin, suggesting wide tropism. The prevalence of HBLV antibody in the normal population was 26% though clear differences between different populations were observed. The prevalence of HBLV antibody an elevated antibody titer was higher in sera from certain malignancies, Sjögrens syndrome and sarcoidosis. Antibody to HBLV was also elevated in AIDS patients and patients with chronic fatigue syndrome. HBLV-DNA was detected in some B-cell lymphomas. The broad in vitro tropism, combined with immunological and molecular evidence of HBLV infection in individuals raise the question of the pathogenicity of this virus in some diseases. Because in vitro co-infection of CD4 cells by HBLV and HIV leads to enhanced degeneration, this raises the possibility that infection in AIDS patients by both viruses can aggravate the HIV-induced immunodeficiency. Specific reagents and immunological and molecular assays are currently being investigated, which will aid in virus detection in cells from patients, and in elucidating the possible pathogenesis of HBLV.

Confocal light absorption and scattering spectroscopic microscopy monitors organelles in live cells with no exogenous labels

This article reports the development of an optical imaging technique, confocal light absorption and scattering spectroscopic (CLASS) microscopy, capable of noninvasively determining the dimensions and other physical properties of single subcellular organelles. CLASS microscopy combines the principles of light-scattering spectroscopy (LSS) with confocal microscopy. LSS is an optical technique that relates the spectroscopic properties of light elastically scattered by small particles to their size, refractive index, and shape. The multispectral nature of LSS enables it to measure internal cell structures much smaller than the diffraction limit without damaging the cell or requiring exogenous markers, which could affect cell function. Scanning the confocal volume across the sample creates an image. CLASS microscopy approaches the accuracy of electron microscopy but is nondestructive and does not require the contrast agents common to optical microscopy. It provides unique capabilities to study functions of viable cells, which are beyond the capabilities of other techniques.

Clinical characterization and outcomes of preeclampsia with normal angiogenic profile

Objective: To compare the clinical characteristics and outcomes of preeclamptic women presenting with a normal plasma angiogenic profile with those subjects who are characterized by an abnormal angiogenic profile. Methods: This was a secondary analysis of a prospective cohort study in women presenting to obstetrical triage at <37 weeks of gestation and diagnosed with preeclampsia within 2 weeks of enrollment and in whom angiogenic factors (sFlt1 and PlGF) measurements were available. Patients were divided into two groups based on their circulating levels of these factors described as a ratio; the sFlt1/PlGF ratio, non-angiogenic preeclampsia (sFlt1/PlGF ratio <85) and angiogenic preeclampsia (sFlt1/PlGF ratio ≥85). The data are presented by sFlt1/PlGF category using median and quartile 1–quartile 3 for continuous variables and by frequency and sample sizes for categorical variables. Results: In our cohort, the patients with non-angiogenic preeclampsia (N = 46) were more obese [BMI: 35.2 (31.6, 38.7) versus 31.1 (28.0, 39.0), p = 0.04], more likely to have preexisting diabetes (21.7% versus 2.0%, p = 0.002) and presented at a later gestational age [35 (32, 37) versus 32 (29, 34) weeks, p < 0.0001] as compared with women with angiogenic preeclampsia (N = 51). Women with non-angiogenic preeclampsia had no serious adverse outcomes (elevated liver function tests/low platelets: 0% versus 23.5%, abruption: 0% versus 9.8%, pulmonary edema: 0% versus 3.9%, eclampsia: 0% versus 2.0 %, small for gestational age: 0% versus 17.7% and fetal/neonatal death: 0% versus 5.9%) as compared with women with angiogenic preeclampsia. The rate of preterm delivery <34 weeks was 8.7% in non-angiogenic preeclampsia compared with 64.7% in angiogenic preeclampsia (p < 0.0001). Interestingly, delivery between 34 and 37 weeks and resource utilization (hospital admission days) were similar in the two groups. Conclusion: In contrast to the angiogenic form, the non-angiogenic form of preeclampsia is characterized by little to no risk of preeclampsia-related adverse outcomes, other than iatrogenic prematurity. Incorporation of angiogenic biomarkers in the evaluation of preeclampsia may allow accurate and early identification of severe disease.

Polymerase chain reaction amplification and in situ hybridization for the detection of human B-lymphotropic virus

Polymerase chain reaction amplification (PCR) is a recently described technique that allows for the amplification of a given sequence of DNA. It can be used to reliably amplify sequences of up to 3 kb within hours. The amplified sequence can then be recognized by hybridization with a specific probe after transfer onto nitrocellulose or nylon paper. We used PCR to recognize human B-lymphotropic virus (HBLV or HHV-6) specific sequences in various tumors as well as in the blood of patients with AIDS. Sixty-three specimens of DNA extracted from peripheral blood of patients with AIDS as well as DNA extracted from various lymphoproliferative disorders were analysed; 52 out of 63 (83%) patients with AIDS were found to have amplification of the HHV-6 specific sequence; 2 out of the 63 (3%) had equivocal amplification and 9 (14%) were found to be negative. Twenty out of 23 tumors were found to have amplified HBLV-specific sequences. Only one of these tumors was positive by Southern hybridization on restriction enzyme digested genomic DNA. In situ hybridization of clinical specimens using radiolabelled RNA probes or hapten-labelled DNA probes was used to detect the presence of HBLV in tumors. Three tumors of B cell origin were found to be positive for HBLV.

Confocal light absorption and scattering spectroscopic microscopy

We have developed a novel optical method for observing submicrometer intracellular structures in living cells, which is called confocal light absorption and scattering spectroscopic (CLASS) microscopy. It combines confocal microscopy, a well-established high-resolution microscopic technique, with light-scattering spectroscopy. CLASS microscopy requires no exogenous labels and is capable of imaging and continuously monitoring individual viable cells, enabling the observation of cell and organelle functioning at scales of the order of 100 nm.

Virologic studies in a case of transfusion-associated AIDS.

The authors present the case of a 60-year old woman who developed acquired immunedeficiency syndrome (AIDS) 43 months after a blood transfusion. The blood donor was identified as a sexually active homosexual man who frequently used intravenous drugs. Although he was asymptomatic, physical examination revealed generalized lymphadenopathy of cervical, axillary, and inguinal areas. 47 months after his blood donation, the donor underwent lymph node biopsy which revealed follicular hyperplasia. He was seropositive for antibodies to human T-lymphotropic virus type III (HTLV-III) and had serologic evidence of prior infection with cytomegalovirus, Epstein-Barr virus, and hepatitis B virus. On the other hand, he had a normal total lymphocyte count and normal T-cell subsets. Striking was the difference between the seropositivity of the high-risk donor and the seronegativity of the patient with transfusion-associated AIDS with respect to cytomegalovirus, Epstein-Barr virus, and hepatitis B virus. The demonstration of apparent transmission of HTLV-III by blood transfusion and the inability to detect evidence of infection with other viruses strongly indicate a primary etiologic role for this virus in the pathogenesis of AIDS. The high-risk blood donor continues to be an asymptomatic carrier of HTLV-III. The authors are currently comparing the HTLV-III isolates obtained from the blood donor with the HTLV-III-related RNA sequences detected in the spleen of the patient in order to determine whether molecular changes occurred after transmission of the virus that might be related to the development of AIDS.

Antibody prevalence to HBLV (human herpesvirus-6, HHV-6) and suggestive pathogenicity in the general population and in patients with immune deficiency syndromes

Detailed serologic screening showed an antibody prevalence to HBLV (HHV-6) in the general population of 26% if very strict criteria for antibody positivity were applied. Lower and borderline antibody titers yet may be found in up to 63% of the population. Only 17% of these persons have clinical symptoms; in the majority infection remains silent. HHV-6 infection apparently occurs already quite early in life, and initial symptoms can occur, such as short-term high fever, sore throat, local lymphadenopathy and skin rash. Lesions disappear without specific treatment. The frequency of positive antibody tests at higher titers rises in patients with immune deficiency and with atypical lymphoproliferative diseases to 60 and 75%. The rise in antibody titers is associated in patients with immune deficiency by characteristic shifts of blood lymphocyte populations, essentially by increase in immature T-lymphocytes. Highest titers are found in patients with lymphoproliferative syndromes, yet the percentage of atypical lymphoid cells harboring the viral genome is low (about 2% of seropositive patients). Thus it appears, that HBLV, similar to other herpesviruses such as Epstein-Barr virus, usually causes a silent seroconversion, yet may be associated with variable clinical pathology when persisting in an active state. Its pathogenic effect might be rather a cofactor contributing to immune disturbance than overt oncogenicity.

The use of angiogenic biomarkers to differentiate non-HELLP related thrombocytopenia from HELLP syndrome

OBJECTIVE Preeclampsia (PE) is diagnosed using clinical criteria and in atypical cases the diagnosis may be inaccurate as there are no specific tests to confirm or exclude PE. This study sought to evaluate the utility of angiogenic biomarkers, sFlt1, sEng and PlGF to distinguish patients with gestational thrombocytopenia and immune thrombocytopenic purpura (ITP) from patients with thrombocytopenia resulting from the HELLP (hemolysis, elevated liver enzymes and low platelets) syndrome, a complication of severe PE. METHODS Serum was collected and the angiogenic biomarkers of patients with ITP and gestational thrombocytopenia (N = 9) were compared to patients with HELLP (N = 11) and PE (N = 11). Circulating levels of these angiogenic biomarkers were also compared by gestational age to 1564 randomly selected normotensive women from the Calcium for Preeclampsia Prevention study. RESULTS Patients with non-HELLP thrombocytopenia had lower sFlt1 (7.3 +/- 3.8 ng/ml vs. 15.5 +/- 5 ng/ml, P < 0.001), lower sEng (8.7 +/- 3.6 vs. 34 +/- 17, P < 0.001) and higher PlGF (484 +/- 412 vs. 66.3 +/- 44, P = 0.003) than patients with HELLP syndrome. Angiogenic factor abnormalities in patients with PE were similar to patients with HELLP syndrome, suggesting a common pathogenesis. Patients with non-HELLP thrombocytopenia had angiogenic profiles similar to normotensive controls, whereas patients with HELLP syndrome had levels higher than the 90th percentile for sFlt1 and sEng and lower than the 10th percentile for PlGF. CONCLUSIONS Angiogenic biomarkers may be useful in excluding conditions that mimic PE.