The economic impact of implementing a multiple inflammatory biomarker-based approach to identify, treat, and reduce cardiovascular risk.

OBJECTIVES: To develop an economic model to estimate the change in the number of events and costs of non-fatal myocardial infarction (MI) and non-fatal ischemic stroke (IS) as a result of implementing routine risk-stratification with a multiple inflammatory biomarker approach. METHODS: Reductions in the numbers of non-fatal MI and non-fatal IS events and in related per-member-per-month (PMPM) and 5-year costs (excluding test costs) due to biomarker testing were modeled for a US health plan with one million beneficiaries. Inputs for the model included literature-based MI and IS incidence rates, healthcare costs associated with MI and IS, laboratory results of biomarker testing, MI and IS hazard ratios related to biomarker levels, patient monitoring and intervention costs and use/costs of preventative pharmacotherapy. Preventative pharmacotherapy inputs were based on an analysis of pharmacy claims data. Costs savings (2013 USD) were assessed for patients undergoing biomarker testing compared to the standard of care. Data from MDVIP and Cleveland Heart Lab supported two critical inputs: (1) treatment success rates and (2) the population distribution of biomarker testing. Incidence rates, hazard ratios, and other healthcare costs were obtained from the literature. RESULTS: For a health plan with one million members, an estimated 21,104 MI and 22,589 IS events occurred in a 5-year period. Routine biomarker testing among a sub-group of beneficiaries ≥35 years old reduced non-fatal MI and IS events by 2039 and 1869, respectively, yielding cost savings of over $187 million over 5 years ($3.13 PMPM), excluding test costs. Results were sensitive to changes in treatment response rates. Nonetheless, cost savings were observed for all input values. CONCLUSIONS: This study suggests that health plans can realize substantial cost savings by preventing non-fatal MI and IS events after implementation of routine biomarker testing. Five-year cost savings before test costs could exceed $3.13 PMPM.

Characterization of high-risk HIV-1 seronegative hemophiliacs.

Mechanisms that protect most high-risk HIV-1 seronegative (HRSN) persons are not well understood. Among hemophiliacs from the Multicenter Hemophilia Cohort Study who remained HIV-1 seronegative despite a high (94%) risk for acquisition of HIV-1 infection, only 7/43 were homozygous for the protective CCR5 Delta32 polymorphism. Among the remainder, neither CCR5 density nor beta-chemokine production, nor in vitro susceptibility to infection with the HIV-1 isolate JR-FL could distinguish HRSN hemophiliacs from healthy controls. When compared to lymphocytes of healthy controls not at risk for HIV-1 infection, diminished spontaneous lymphocyte proliferation was seen in lymphocytes of HRSN hemophiliacs as well as in lymphocytes of hemophiliacs not at risk for HIV-1 infection. Surprisingly sera/plasmas obtained from high-risk HIV-1 seropositve hemophiliacs prior to seroconversion more often contained alloreactive antibodies than date-matched sera/plasmas obtained from HRSN hemophiliacs. Thus alloreactivity may predispose to acquisition of HIV-1 infection after parenteral exposure.

Effects of Mycobacterium avium complex-infection treatment on cytokine expression in human immunodeficiency virus-infected persons: results of AIDS clinical trials group protocol 853.

Human immunodeficiency virus (HIV) type 1-infected persons with newly diagnosed Mycobacterium avium complex (MAC) bacteremia were enrolled in an 8-week study to determine whether treatment of MAC infection is associated with decreases in plasma tumor necrosis factor (TNF)-alpha levels. Blood specimens were obtained for quantitative MAC cultures and to determine plasma levels of HIV RNA, TNF-alpha, and other proinflammatory cytokines. MAC levels decreased by 1.75 log at week 4 (P=.008) and by 2.48 log at week 8 (P=.001). Plasma TNF-alpha decreased by 0.15 log at week 4 (P=.042) and by 0. 40 log at week 8 (P=.027). Plasma interleukin (IL)-6 decreased by 0. 56 log at week 8 (P=.039). There were nonsignificant trends (P<.10) for plasma levels of IL-1beta and HIV RNA to decrease at week 8. Nonsignificant decreases in plasma levels of TNF-alpha, IL-1beta, IL-6, and HIV RNA were also seen in those individuals who remained on stable antiretroviral therapy throughout the 8 weeks of the study.

Lymphocyte proliferation assays may underestimate antigen responsiveness in human immunodeficiency virus infection.

The objective of this study was to examine the relationships among lymphocyte proliferation, interferon-gamma (IFN-gamma) production, and apoptosis in peripheral blood mononuclear cells (PBMC) of HIV-1-infected patients and controls. PBMC were prepared from 19 HIV-1-infected patients and 16 healthy controls. Using tetanus toxoid (TT) as a recall antigen, we assessed lymphocyte proliferation using [(3)H]thymidine incorporation after 2, 4, 6, and 7 days' culture and IFN-gamma production in 48-h culture supernatants by ELISA. Apoptosis was measured using TdT-mediated dUTP nick-end labeling. Median stimulation indices (SIs) in HIV-1-infected patients were 2.8 and 3.7 as opposed to 24.9 and 25.1 in healthy controls after 6 and 7 days' culture, respectively (P < 0. 001). Among the controls, peak proliferation was seen after 7 days in culture whereas in patients, SIs peaked at 4 days and fell progressively by days 6 and 7. At 2 and 4 days of stimulation with tetanus, patients' T cells showed increased apoptosis (19 and 25%) vs 12 and 15% apoptosis seen in controls' cells, P < 0.05. Interferon-gamma in 48-h supernatants of TT-stimulated PBMC was comparable among patients and controls. Whereas in our system, 6 and 7 day assays of lymphocyte proliferation provide increasing responses to TT among healthy controls, these durations of culture may underestimate antigen responsiveness in HIV-1 infection. Cell death due to apoptosis may account for this phenomenon. Whether shorter term or longer term assays of lymphocyte responsiveness more accurately reflect in vivo immune competence is unknown. Nonetheless, shorter duration assays may provide a more realistic estimate of the frequency of antigen-reactive cells in persons with HIV-1 infection.

HIV infection perturbs DNA content of lymphoid cells: partial correction after 'suppression' of virus replication.

OBJECTIVE: To examine the DNA content of circulating lymphocytes obtained from HIV-1-infected persons and to explore the effects of antiretroviral therapy on these indices. DESIGN: Cross-sectional analysis and 48-week open label treatment trial (AIDS Clinical Trials Group Protocol 315) of zidovudine, lamivudine and ritonavir. METHODS: Peripheral blood lymphocytes were obtained from HIV-1-infected patients and healthy controls and after 48 h of in vitro cultivation were stained with propidium iodide and analyzed for DNA content by flow cytometry. RESULTS: HIV-1-infected patients had more hypodiploid cells (19%), fewer G0-G1 phase cells (70%) and more S phase cells (10%) than did healthy controls (8%, 85% and 5% respectively; P = 0.002). Patients with sustained suppression of plasma HIV-1 RNA levels after antiretroviral therapy had only modest improvements in these indices. In contrast, patients who failed to suppress plasma HIV-1 RNA levels had decreases in G0-G1 cells to 54% (P = 0.032) and increases in S phase cells to 24% (P = 0.055). Plasma HIV-1 RNA levels and the percentage of S phase cells were correlated (r, 0.23; P = 0.047). In patients failing antiretroviral therapy, there was an inverse correlation between the percentage of G0-G1 cells and expression of the activation antigens CD38 and HLA-DR on CD4 cells (r, -0.409; P = 0.016) and CD8 cells (r, -0.363; P = 0.035). CONCLUSIONS: Lymphocytes obtained from HIV-1-infected patients display perturbations in DNA content after brief cultivation in vitro reflective of immune activation in vivo. The marginal improvement in these indices after 'successful' suppression of HIV-1 replication suggests that even low levels of HIV-1 replication are sufficient to induce immune activation and perturbations in DNA content.

Levels of proinflammatory cytokines in plasma after pneumoccoccal immunization in human immunodeficiency virus type 1-infected patients.

To ascertain if immunization with pneumococcal polysaccharide vaccine is associated with rises in the levels of proinflammatory cytokines in the plasma of human immunodeficiency virus type 1 (HIV-1)-infected patients, the levels of tumor necrosis factor alpha (TNF-alpha) and interleukin-6 (IL-6) were measured serially after immunization. IL-6 levels rose an average of 2.2- and 2.1-fold 6 and 8 h after immunization, respectively, but TNF-alpha levels remained unchanged. The levels of these cytokines were stable in unimmunized controls. Immunization with pneumococcal polysaccharide vaccine induces increases in the levels of IL-6 in the plasma of persons with HIV-1 infection.

No evidence of KSHV/HHV-8 in mycosis fungoides or associated disorders.

The recently discovered human virus known as Kaposi's sarcoma-associated herpesvirus (KSHV) or human herpesvirus-8 (HHV-8) has been associated with body-cavity-based lymphomas in AIDS patients. It is most closely related to two other herpesviruses, the Epstein-Barr virus and herpesvirus saimiri, which are known to be associated with lymphomas in humans and nonhuman primates, respectively. To determine whether KSHV/HHV-8 is involved in the pathogenesis of mycosis fungoides (MF) and related disorders, we used a genomic PCR assay followed by confirmatory Southern blot analysis with a nested oligonucleotide probe to analyze cases for the presence of this virus. The specimens studied included fresh-frozen lesional tissues obtained from 16 patients with MF, seven with lymphomatoid papulosis, seven with primary cutaneous CD30+ large cell lymphoma of T-cell lineage, and five with Hodgkin's disease. Two T-cell tumor lines were also studied: MT4 (derived from a patient with adult T-cell leukemia/lymphoma) and Jurkat (derived from a patient with T-cell acute lymphoblastic leukemia). All cases were uniformly negative for KSHV/HHV-8, whereas Kaposi's sarcoma-positive controls and human beta-globin DNA integrity controls were appropriately positive. These findings provide strong evidence against a role for KSHV/HHV-8 in the pathogenesis of MF or associated lymphoproliferative disorders.

High incidence of Kaposi's sarcoma-associated herpesvirus and Epstein-Barr virus in tumor lesions and peripheral blood mononuclear cells from patients with Kaposi's sarcoma in Uganda.

With the advent of AIDS, Kaposi's sarcoma (KS) has become one of the leading malignancies in sub-Saharan Africa. Recently, DNA sequences from a new human herpesvirus called KS-associated herpesvirus (KSHV) or human herpesvirus type 8 have been found in KS tumor lesions in high frequency. Analyses of tumor lesions from 38 Ugandan KS patients indicated a uniform presence of KSHV in KS tumor lesions as revealed by polymerase chain reaction and Southern hybridization. In contrast, only 31% (11/36) of the normal skin biopsies from the same patient population were positive. The frequency of KSHV DNA detection in peripheral blood mononuclear cells (PBMC) of KS patients was also high (84%, 31/37). Similar analyses revealed the presence of cytomegalovirus (21% in KS lesions) to be discordant with KS development. A large number of KS lesions (87%, 33/38) and KS PBMC (70%, 26/37) were, however, positive for Epstein-Barr virus sequences. In addition, KSHV DNA was not found in the PBMC of Ugandans without KS.

Kaposi's Sarcoma in AIDS: High incidence of Kaposi's sarcoma-associated herpesvirus and Epstein-Barr virus in tumor lesions and peripheral blood mononuclear cells from patients with Kaposi's sarcoma in Uganda

With the advent of AIDS; Kaposi's sarcom (KS) has become one of the leading malignancies in sub-Saharan Africa. Recently; DNA sequences from a new human herpesvirus called KS-associated herpesvirus (KSHV) or human herpesvirus type 8 have been found in KS tumor lesions as revealed by polymerase chain reaction and Southern hybridization. In contrast; only 31(11/36) of the normal skin biopsies from the same patient population were positive. The frequency of KSHV DNA detection in peripheral blood mononuclear cells (PBMC) of KS patients was also high (84; 31/37). Similar analyses revealed the presence of cytomegalovirus (21in KS lesions) to be discordant with KS PBMC (70; 26/37) were; however; positive for Epstein-Barr virus sequences. In addition; KSHV DNA was not found in the PBMC of Ugandans without KS. (Source: J. Infect. Dis. 1997 April; 175(4):947-50)

HIV type 1 Tat protein induces apoptosis and death in Jurkat cells.

Jurkat cells stably expressing high levels of the HIV-1 Tat protein were generated after transfection with an Epstein-Barr virus-based episomal replicon and selection in hygromycin B. The Jurkat Tat transfectants exhibited a longer doubling time when compared to Jurkat cells or Jurkat cells transfected with the control parent plasmid. Cell cycle analysis revealed comparable durations of each phase of the cell cycle in the Tat and control transfectants. Flow cytometric analysis using Hoechst 33342 and propidium iodide staining revealed that the Tat transfectants exhibited a higher percentage of apoptotic cells when compared to the control transfectants (29.1 +/- 3.1 vs. 11.43 +/- 3.1%). Incubation of Jurkat cells with recombinant HIV-1 Tat protein resulted in induction of apoptosis. The HIV-1 Tat protein induces apoptosis in a CD4-positive T cell line. Tat-induced programmed cell death may contribute to the lymphocyte depletion seen in persons infected with HIV-1.

Postnatal support. Listening: an under-valued health visiting skill.

The experience of being listened to can be a turning point in people's lives, particularly in the early months of parenthood, write Shona Reed-Purvis and Sally Dakin. This is a time when health visitors are ideally placed to offer emotional support. But training may be necessary to help the practitioner move beyond the traditional directive role engendered by old-style nurse training and expected by many clients, and to encourage the client to take control. Here they describe a training programme developed at the request of Oxford City health visitors to help them use their skills more effectively.

Suppression of interleukin-2 and interleukin-2 receptor expression in Jurkat cells stably expressing the human immunodeficiency virus Tat protein.

The Jurkat T cell line was stably transfected with an Epstein-Barr virus-based episomal replicon designed to express high levels of the HIV-1 Tat protein. After selection in hygromycin B, high-level Tat activity was detected in 3 of 18 transfected cell lines. After stimulation with phytohemagglutinin (PHA) and phorbol myristate acetate (PMA), Tat transfectants with high Tat expression showed diminished expression of interleukin-2 (IL-2) and the interleukin-2 receptor alpha chain (IL-2R) when compared to untransfected Jurkat cells or Jurkat cell lines transfected with the parent control plasmid. Sublines derived from the high-level Tat transfectants with reduced Tat activity showed normalization of PHA/PMA-induced IL-2 expression. Northern analysis showed diminished expression of IL-2 and IL-2R mRNA in the stimulated Tat transfectants. Inhibition of IL-2 and IL-2R expression by the HIV-1 Tat protein may contribute to the immune suppression that characterizes HIV-1 infection.

Fetal hemoglobin levels in sickle cell disease and normal individuals are partially controlled by an X-linked gene located at Xp22.2.

Fetal hemoglobin (Hb F) production in sickle cell (SS) disease and in normal individuals varies over a 20-fold range and is under genetic control. Previous studies suggested that variant Hb F levels might be controlled by genetic loci separate from the beta-globin complex on chromosome 11. Using microscopic radial immunodiffusion and flow cytometric immunofluorescent assays to determine the percentage of F reticulocytes and F cells in SS and nonanemic individuals, we observed that F-cell levels were significantly higher in nonanemic females than males (mean +/- SD, 3.8% +/- 3.2% v 2.7% +/- 2.3%). F-cell production as determined by F reticulocyte levels in SS females was also higher than in SS males (17% +/- 10% v 13% +/- 8%). We tested the hypothesis that F-cell production in both normal and anemic SS individuals was controlled by an X-linked locus with two alleles, high (H) and low (L). Using an algorithm to determine the 99.8% confidence interval of a normal distribution in nonanemic individuals, we estimated that males and females with at least one H allele had greater than 3.3% F cells. Comparisons of male-male or female-female SS sib pairs with discordant F reticulocyte levels distinguished two phenotypes in SS males (L, less than 12%; H, greater than 12%) and three phenotypes in SS females (LL, less than 12%; HL, 12% to 24%, HH greater than 24%). Linkage analysis using polymorphic restriction sites along the X chromosome in eight SS and one AA family localized the F-cell production (FCP) locus to Xp22.2, with a maximum lod score (logarithm of odds of linkage v independent assortment) of 4.6 at a recombination fraction of 0.04.

Heightened complement sensitivity of acquired immunodeficiency syndrome lymphocytes related to diminished expression of decay-accelerating factor.

Although the human immunodeficiency virus can induce cytopathic changes in human lymphocytes in vitro, the mechanism(s) underlying progressive lymphopenia in patients with AIDS and AIDS-related complex has not been elucidated. To investigate this issue, peripheral blood lymphocytes of AIDS and AIDS-related complex patients and healthy control subjects were examined for their ability to resist homologous complement-mediated lysis. Upon sensitization with monoclonal antibodies to major histocompatibility complex class I antigen, as much as 48% lysis of patients' cells was observed in as little as a 1:32 dilution of human serum compared to 18 +/- 8% (mean +/- SD) lysis of controls' cells even in a 1:8 dilution of human serum. To investigate the mechanism of the abnormal complement sensitivity, AIDS and AIDS-related complex cells were analyzed for expression of decay-accelerating factor (DAF), a complement regulatory protein that functions intrinsically in blood cell membranes to prevent complement activation on their surfaces. Flow cytometric assays using anti-DAF monoclonal antibodies demonstrated that patients' lymphocytes and monocytes were DAF-deficient, in contrast to their polymorphonuclear leukocytes, which showed normal DAF levels. Expression of DAF was diminished on CD4+ as well as CD8+ T-lymphocyte subpopulations as opposed to expression of CD3, which was comparable in patients and controls. Incubation of normal lymphocytes with anti-DAF monoclonal antibodies or phosphatidylinositol-specific phospholipase C, an enzyme that cleaves DAF, enhanced lysis. Conversely, reconstitution of patients' cells with exogenous DAF reduced their lysis. The findings of heightened complement sensitivity and DAF deficiency of patients' lymphocytes in vitro suggest the possibility that the DAF deficit may contribute to the progressive lymphopenia of AIDS in vivo.

Regulation of human T-cell production of interleukin 2 by Leu 11 (CD16) positive large granular lymphocytes.

Peripheral blood large granular lymphocytes (LGL) expressing Leu 11 (CD16) antigen with potent natural killer cytotoxicity inhibited soluble antigen-induced T-cell production of interleukin 2 (IL-2). Depletion of Leu 11-reactive cells from T-cells doubled IL-2 activity (P less than 0.05). Leu 11-enriched cells did not express high affinity IL-2 receptors nor did they deplete IL-2 activity from culture media. Upon addition in low ratios to Leu 11-depleted cells, Leu 11-enriched fractions increased antigen-induced IL-2 production three-fold (P less than 0.05), whereas at higher ratios IL-2 production was suppressed P less than 0.01. Additionally, adherent monocytes were increasingly accessory when added in graded numbers to Leu 11-depleted but not T-cell cultures. In the presence of small numbers (5%) of Leu 11-enriched cells, however, monocytes down-regulated IL-2 production of Leu 11-depleted cell cultures. Thus Leu 11-reactive lymphocytes have noncytotoxic functions and may play a major role in immunoregulation.

Comparison of magnetic resonance imaging with somatosensory testing in MS suspects.

Two hundred patients suspected of having multiple sclerosis (MS), including 42 with progressive myelopathy and 11 with optic neuritis, were investigated with somatosensory evoked potentials (SEPs) and magnetic resonance imaging (MRI). Most had minimal neurological deficit, the mean Kurtzke scale being 2.65. There were 117 patients had two or more MRI lesions suggestive of MS, with a total of 527 lesions identified; 290 (55%) involved the somatosensory pathways, most commonly lying in the mid-periventricular region (thalamo-cortical radiations). There was good correlation between positive and negative MRIs and SEPs. The MRI was abnormal more frequently than the SEP, except in progressive myelopathy when both were abnormal with equal frequency. It is proposed that some cases of myelopathy in MS may be due to periventricular rather than spinal pathology. The morphology of the MRI lesion would favor conduction block not slowing of the SEP as being the prime abnormality. This appeared to be true of leg SEPs.

Isolation of heavy chain isoenzymes of myosin subfragment 1 by high performance ion exchange chromatography.

The procedure of high performance ion-exchange chromatography has been used to fractionate subfragment 1 of myosin (SF1) into its isoenzymic forms. In contrast to conventional ion-exchange procedures which yield two fractions corresponding to SF1(A1) and SF1(A2), the high performance liquid chromatography (HPLC) procedure resolves SF1 into four discrete fractions. The first pair that is eluted appears to be A1-containing isoenzymes while the latter pair corresponds to the A2 forms based on their polypeptide compositions by gel electrophoresis in the presence of sodium dodecyl sulfate. By gel electrophoresis under nondenaturing conditions it is not possible to differentiate between the two fractions corresponding to each isoenzyme. Although very minor differences between the fractions can be seen by the presence of extraneous peptides, these are present in far below stoichiometric amounts and, therefore, make it very unlikely that the superior fractionation by the HPLC procedure is based on their presence. An examination of the heavy chain heterogeneity in each of these fractions by peptide mapping revealed that the extra separation was based on this factor. Thus the HPLC procedure is capable of providing separation of SF1 into heavy chain-based isozymes as well as the light chain forms. ATPase measurements of these fractions reveal only minor differences in the Ca2+- and EDTA-activated ATPase.