Tzong-Hae Lee

Maternal alloantigens promote the development of tolerogenic fetal regulatory T cells in utero.

As the immune system develops, T cells are selected or regulated to become tolerant of self antigens and reactive against foreign antigens. In mice, the induction of such tolerance is thought to be attributable to the deletion of self-reactive cells. Here, we show that the human fetal immune system takes advantage of an additional mechanism: the generation of regulatory T cells (Tregs) that suppress fetal immune responses. We find that substantial numbers of maternal cells cross the placenta to reside in fetal lymph nodes, inducing the development of CD4+CD25highFoxP3+ Tregs that suppress fetal antimaternal immunity and persist at least until early adulthood. These findings reveal a form of antigen-specific tolerance in humans, induced in utero and probably active in regulating immune responses after birth.

Kinetics of fetal cellular and cell‐free DNA in the maternal circulation during and after pregnancy: implications for noninvasive prenatal diagnosis

BACKGROUND: Fetal genetic material is detectable in the maternal circulation and has been used for noninvasive prenatal diagnosis. However, few data are available concerning its quantity and natural history during gestation.

Quantitation of genomic DNA in plasma and serum samples: higher concentrations of genomic DNA found in serum than in plasma.

BACKGROUND: Plasma and serum samples have been used to detect cell‐free genomic DNA in serum or plasma in certain pathologic conditions such as systemic lupus erythematosus, pulmonary embolism, and malignancies, as well as in fetal cell chimerisms in maternal serum and/or plasma. In this study, baseline concentrations of cell‐free DNA in serum and plasma samples were evaluated for the study of posttransfusion chimerism.

Evidence for Persistent Low-Level Viremia in Individuals Who Control Human Immunodeficiency Virus in the Absence of Antiretroviral Therapy

ABSTRACT A subset of antiretroviral-untreated, human immunodeficiency virus (HIV)-infected individuals are able to maintain undetectable plasma HIV RNA levels in the absence of antiretroviral therapy. These “elite” controllers are of high interest as they may provide novel insights regarding host mechanisms of virus control. The degree to which these individuals have residual plasma viremia has not been well defined. We performed a longitudinal study of 46 elite controllers, defined as HIV-seropositive, antiretroviral-untreated individuals with plasma HIV RNA levels of <50 to 75 copies/ml. The median duration of HIV diagnosis was 13 years, the median baseline CD4+ T-cell count was 753 cells/mm3, and the median duration of follow-up was 16 months. Plasma and cellular HIV RNA levels were measured using the transcription-mediated amplification (TMA) assay (estimated limit of detection of <3.5 copies RNA/ml). A total of 1,117 TMA assays were performed (median of five time points/subject and four replicates/time point). All but one subject had detectable plasma HIV RNA on at least one time point, and 15 (33%) subjects had detectable RNA at all time points. The majority of controllers also had detectable cell-associated RNA and proviral DNA. A mixed-effect linear model showed no strong evidence of change in plasma RNA levels over time. In conclusion, the vast majority (98%) of elite controllers had measurable plasma HIV RNA, often at levels higher than that observed in antiretroviral-treated patients. This confirms the failure to eradicate the virus, even in these unique individuals who are able to reduce plasma viremia to very low levels without antiretroviral therapy.

Antiretroviral Therapy Initiated Within 6 Months of HIV Infection Is Associated With Lower T-Cell Activation and Smaller HIV Reservoir Size

Background. CD4(+)/CD8(+) T-cell activation levels often remain elevated in chronic human immunodeficiency virus (HIV) infection despite initiation of antiretroviral therapy (ART). T-cell activation predicts early death and blunted CD4+ T-cell recovery during ART and may affect persistent HIV reservoir size. We investigated whether very early ART initiation is associated with lower on-therapy immune activation and HIV persistence. Methods. From a cohort of patients with early HIV infection (<6 months duration since infection) we identified persons who started ART early (<6 months after infection) or later (≥2 years after infection) and maintained ≥2 years of virologic suppression; at-risk HIV-negative persons were controls. We measured CD4(+)/CD8(+) T-cell activation (percent CD38(+)/HLA-DR(+)) and HIV reservoir size (based on HIV DNA and cell-associated RNA levels). Results. In unadjusted analyses, early ART predicted lower on-therapy CD8(+) T-cell activation (n = 34; mean, 22.1%) than achieved with later ART (n = 32; mean, 28.8%; P = .009), although levels in early ART remained elevated relative to HIV-negative controls (P = .02). Early ART also predicted lower CD4+ T-cell activation than with later ART (5.3% vs 7.5%; P = .06). Early ART predicted 4.8-fold lower DNA levels than achieved with later ART (P = .005), and lower cell-associated RNA levels (difference in signal-to-cutoff ratio (S/Co), 3.2; P = .035). Conclusions. ART initiation <6 months after infection is associated with lower levels of T-cell activation and smaller HIV DNA and RNA reservoir size during long-term therapy.

Bacillary Angiomatosis and Bacillary Splenitis in Immunocompetent Adults

Bacillary angiomatosis and parenchymal bacillary peliosis are recently described vascular disorders associated with infection by Rochalimaea henselae and Rochalimaea quintana, which occur in patients with either human immunodeficiency virus (HIV) infection or drug-induced immune suppression [1-5]. In addition, R. henselae and R. quintana (also the agent of trench fever) are both members of the family Rickettsiaceae [1, 4, 5], and infections due to Rochalimaea species have been associated with exposure both to cats [2] and to arthropod vectors [1, 5]. We describe five patients with cutaneous bacillary angiomatosis or bacillary splenitis without evidence of HIV infection who were determined to be immunocompetent after immunologic evaluation. In three patients with both cat and cat flea exposures, infection by R. henselae was confirmed by amplification and sequencing of 16S rDNA from an infected tissue specimen. Methods Four patients with characteristic vascular lesions of cutaneous bacillary angiomatosis [1, 3] were examined. Patient 2, who lacked vascular lesions, had a 2-week history of left-sided abdominal pain, shortness of breath, low-grade fever, nausea, diarrhea, and weight loss of 3.5 kg. Results of laboratory studies included hematocrit, 0.21 (2 months before it had been 0.34); lactate dehydrogenase, 1170 U/L; total bilirubin, 53 mol/L (3.1 mg/dL); conjugated bilirubin, 24 mol/L (1.4 mg/dL); and mildly elevated hepatic transaminases and alkaline phosphatase. A computed axial tomography scan of the abdomen showed a normal liver and hypersplenism with a 4-cm2 area of focal low attenuation. The patient underwent emergency splenectomy for impending rupture. The 1100-g spleen revealed a 4.5-cm3 mottled area consistent with infarction and two 0.6-cm3 tan nodules beneath the capsular surface. Results of routine bacterial cultures were negative. In June 1991, all five patients had blood drawn for HIV culture, serologic analysis, and polymerase chain reaction studies using techniques described previously [6, 7]. In August 1991, all patients had blood drawn for immunologic evaluation, including quantitative immunoglobulins, complement, lymphocyte subset percentages, neutrophil oxidative burst [8, 9], T-lymphocyte activation studies to phytohemagglutinin, and B-lymphocyte activation studies to pokeweed mitogen [10, 11]. After their blood was drawn, patients had skin test antigens to purified protein derivative, mumps, Trichophyton, and Candida albicans placed and read at 48 hours. The diagnosis of cutaneous bacillary angiomatosis was established using defined histopathologic criteria [3]. Bacterial DNA present in the infected tissue specimen was extracted from either frozen skin biopsy tissue (Patient 4) or from sections of formalin-fixed, paraffin-embedded biopsy specimens [1, 12]. Insufficient tissue was available from Patients 1 and 3. Extracted DNA was amplified by polymerase chain reaction using primers p24E and p12B [1, 12]. Control tissues were simultaneously extracted and amplified [1]. Amplified 16S rDNA products from Patients 2, 4, and 5 were sequenced as described previously [1]. Results Only Patient 2 had a well-documented antecedent chronic illnesshereditary spherocytosis and noninsulin-dependent diabetes mellitusand no patient was receiving immunosuppressive drugs (Table 1). Idiopathic hemochromatosis was diagnosed concomitantly with bacillary angiomatosis in Patient 3. All patients responded to oral antimicrobial therapy of 4 to 6 weeks duration. Histologic examination of skin biopsy specimens from Patients 1, 3, 4, and 5 were diagnostic of bacillary angiomatosis; splenic tissue from Patient 2 showed necrotizing splenitis with fibromyxoid changes, degenerating neutrophils, and mononuclear cells in the absence of both vascular proliferation and granuloma formation. Specimens from all patients showed many bacilli on both Warthin-Starry staining and electron microscopic examination. Table 1. Characteristics of Immunocompetent Patients with Bacillary Angiomatosis and Bacillary Splenitis Amplification of the DNA extracted from infected tissue from Patients 2, 4, and 5 produced a 16S rDNA fragment of approximately 300 base pairs. The amplified DNA fragment from tissue of Patients 2 and 4 is shown in Figure 1. The sequence of this 16S rDNA fragment from Patients 2, 4, and 5 was identical to that of R. henselae [4, 12]. Figure 1. Amplification of Rochalimaea DNA. Rochalimaea henselae R. henselae Discussion We evaluated four patients with cutaneous bacillary angiomatosis and one with bacillary splenitis and found no evidence of HIV infection using a combination of sensitive HIV antibody and antigen assays, as well as viral culture and polymerase chain reaction techniques. In addition, both cellular and humoral immunity were evaluated and no abnormality was found. Because host defense mechanisms against Rochalimaea species are not well understood, defects may have been undetected by our methods. Two patients had underlying illnesses that may be associated with altered immune function, but no consistent immune defect has been associated with these conditions [13-15]. The histopathologic findings were diagnostic for bacillary angiomatosis in all four patients with cutaneous disease; tissue from Patient 2 showed necrotizing splenitis with characteristic bacillary organisms [3, 12]. Amplification and sequencing of rDNA from tissue of Patients 2, 4, and 5 confirmed that the infecting organism was R. henselae. Although we cannot speciate the bacilli seen in abundance with the Warthin-Starry stains of Patients 1 and 3, the histopathologic changes met all criteria for the diagnosis of cutaneous bacillary angiomatosis [3], a disease associated with R. henselae and R. quintana [1]. Bacillary splenitis in the absence of peliosis appears to be another manifestation of R. henselae infection. A clinical spectrum of R. henselae infection may exist, beginning with fever and bacteremia, progressing to bacillary splenitis and finally to bacillary peliosis. Differences in host immune function also may play a role in disease progression. The diagnosis of cutaneous bacillary angiomatosis and bacillary splenitis should be pursued in the immunocompetent patient when clinical or histopathologic features are suggestive. On the basis of these five patients, we recommend treatment with erythromycin or doxycycline for at least 6 weeks when the diagnosis is confirmed.

Cell-Based Measures of Viral Persistence Are Associated With Immune Activation and Programmed Cell Death Protein 1 (PD-1)–Expressing CD4+ T cells

BACKGROUND  Studies aimed at defining the association between host immune responses and human immunodeficiency virus (HIV) persistence during therapy are necessary to develop new strategies for cure. METHODS  We performed a comprehensive assessment of ultrasensitive plasma HIV RNA levels, cell-associated HIV RNA levels, proviral HIV DNA levels, and T cell immunophenotyping in a cohort of 190 subjects in whom HIV levels were suppressed by highly active antiretroviral therapy. RESULTS  The median CD4(+) T cell count was 523 cells/mm(3), and the median duration of viral suppression was 31 months. Cell-associated RNA and proviral DNA levels (but not ultrasensitive plasma HIV RNA levels) were positively correlated with frequencies of CD4(+) and CD8(+) T cells expressing markers of T-cell activation/dysfunction (CD38, HLA-DR, CCR5, and/or programmed cell death protein 1 [PD-1]) (P < .05). Having a low CD4(+) T-cell count despite receipt of virologically suppressive therapy was associated with high cell-associated RNA and proviral DNA levels (P < .01) and higher frequencies of CD4(+) T cells expressing CD38, HLA-DR, CCR5, and/or PD-1 (P < .0001). CONCLUSIONS Cell-based measurements of viral persistence were consistently associated with markers of immune activation and the frequency of PD-1-expressing CD4(+) T cells. Treated patients with a low CD4(+) T-cell count had higher frequencies of PD-1-expressing CD4(+) T cells and cell-based measures of viral persistence, suggesting that HIV infection in these individuals may be more difficult to cure and may require unique interventions.

Dengue viremia in blood donors from Honduras, Brazil, and Australia.

BACKGROUND: Dengue fever and hemorrhagic disease are caused by four dengue virus (DENV) serotypes (DENV‐1 to ‐4), mosquito‐borne flaviviruses with increasing incidence, and expanding global distributions. Documented transfusion transmission of West Nile virus raised concern regarding transfusion‐transmitted DENV.

A Prospective Study of Sexual Transmission of Human T Lymphotropic Virus (HTLV)–I and HTLV-II

BACKGROUND Cross-sectional studies support sexual transmission of human T lymphotropic virus (HTLV)-I/II; however, prospective incidence data, particularly for HTLV-II, are limited. METHODS A cohort of 85 HTLV-positive (30 with HTLV-I and 55 with HTLV-II) blood donors and their stable (>or=6 months) heterosexual sex partners were followed biannually over the course of a 10-year period. RESULTS Four of 85 initially seronegative sex partners of HTLV-I and -II carriers seroconverted, for an incidence rate (IR) of 0.6 transmissions/100 person-years (py) (95% confidence interval [CI], 0.2-1.6). This includes 2 HTLV-I transmissions/219 py (IR, 0.9 transmissions/100 py [95% CI, 0.1-3.3]) and 2 HTLV-II transmissions/411 py (IR, 0.5 transmissions/100 py [95% CI, 0.06-1.8]), with no significant difference by HTLV type. There were 2 male-to-female (IR, 1.2 transmissions/100 py [95% CI, 0.1-4.3]) and 2 female-to-male (IR, 0.4 transmissions/100 py [95% CI, 0.05-1.6) transmissions. HTLV-I or -II proviral load was 2 log10 lower in newly infected partners than in index positive partners who transmitted HTLV (P=.007). CONCLUSIONS The incidence of sexual transmission of HTLV-II may be similar to that of HTLV-I, and female-to-male transmission may play a more important role than previously thought. HTLV-I and -II proviral load may be lower in sexually acquired infection, because of a small infectious dose.

Evidence of persistent low-level viremia in long-term HAART-suppressed, HIV-infected individuals.

Background:HAART can effectively reduce plasma HIV RNA levels to below the level of detection in most HIV-infected patients. The degree to which residual low-level viremia persists during HAART remains unclear. Methods:We identified 180 individuals (median duration of HIV infection 12 years) who had at least two consecutive plasma HIV-1 RNA levels below the level of detection (<50–75 copies/ml) while taking antiretroviral drugs; 36 of 180 had been virologically suppressed for more than 5 years. Longitudinal plasma samples that were taken from these individuals during periods of viral load suppression were selected and analyzed. The isothermal transcription-mediated amplification (TMA) (limit of detection <3.5 copies RNA/ml) assay was used to measure persistent viremia. A ‘detuned’ EIA assay was used to obtain quantitative HIV antibody levels. Results:A total of 1606 TMA assays were performed on 438 specimens in 180 HAART-suppressed individuals (median 3 replicates per specimen). In the first year of viral suppression, plasma RNA levels declined significantly (P = 0.001), but after month 12 there was no evidence for a continued decline (P = 0.383). In the first year of viral suppression, HIV antibody levels also declined (P = 0.054), but after month 12 there was no evidence for a continued decline (P = 0.988). Conclusion:Viremia continued to decline during the first 12 months after viremia became undetectable using conventional methods, and then remained stable. HIV antibody levels also decreased in the first year of viral suppression and then remained stable. Viremia and the HIV-associated host response appear to achieve a steady-state ‘set-point’ during long-term combination therapy.