Fiber technology for reliable repair of skeletal muscle.

Conventional soft-tissue reclosure methods-sutures and staples-require substantial organized-collagen content. Some tissues lack extensive intrinsic collagenous content. Wound disruption consequences range from newly closed abdominal wounds bursting open, to post-cesarean wombs splitting at delivery, to heart valves loosening. Although sutures do approach the theoretical limit of normal force transfer-cross-sectional area times compressive strength, a different paradigm-shear force transfer across the far greater surface attainable by fine fibers parallel to the potential disruptive force could exceed that theoretical limit. Capacity is now the product of frictional coefficient, existing tissue pressure, and contact area. Using a device comprising bundles of poly(ethylene terephthalate) fibers through tissue, we previously coupled muscles to devices and bones. Here we tested an analogous device for reclosing fascia-stripped abdominal wall muscles. In 28 rabbits, fascia-deprived rectus abdominus muscles were reclosed, using the experimental device or conventional sutures. Testing muscles from the 21 three-week survivors, (with closure devices retained-the usual clinical practice) demonstrated experimental failure strength which exceeded that of controls by 58%. Histologically, solid tissue elements did in-grow between fibers for an extensive tissue-prosthetic interface. Both histology and mechanical performance suggest the fiber technology presented herein surpasses conventional sutures in closure of collagen-deficient tissues.

Metabolic abnormalities in HIV type 1-infected children treated and not treated with protease inhibitors.

Our objective was to determine whether HIV-infected children treated with protease inhibitors (PIs) have different blood lipid, insulin, and glucose levels and body composition than HIV-infected children not treated with PIs. A cross-sectional cohort study was performed; in which 23 children were treated with combination antiretroviral therapy including a PI for at least 6 months and 12 children were treated with nucleoside reverse transcriptase inhibitors only (no-PI group). Levels of lipids, apolipoprotein B (apoB), insulin, and glucose were determined in the fasting state. Body composition and fat distribution were determined by anthropometric measurements and dual energy X-ray absorptiometry (DEXA) scan. Total cholesterol levels were higher in the PI-treated children (5.33 +/- 0.87 mM) than in the no-PI children (3.69 +/- 0.59 mM) (p < 0.0001). Similarly, low-density lipoprotein (LDL) levels were also elevated in the PI-treated children (3.27 +/- 0.76 vs. 2.14 +/- 0.51 mM) (p < 0.0001). ApoB and high-density lipoprotein (HDL), and to a lesser degree triglyceride levels, were also increased in the PI-treated children. Apart from percent arm fat as measured by DEXA, there were no differences between the two groups in measures of body composition or in their fasting glucose and insulin levels. The results from this cross-sectional cohort study suggest that the predominant lipid abnormalities associated with treatment with combination antiretroviral therapy including a PI in HIV-1-infected children are elevated total and LDL cholesterol.

Simple, sensitive, and specific detection of human immunodeficiency virus type 1 subtype B DNA in dried blood samples for diagnosis in infants in the field.

The detection of virus is used to diagnose human immunodeficiency virus type 1 (HIV-1) infection in infants due to the persistence of maternal antibodies for a year or more. An HIV-1 DNA PCR assay with simple specimen collection and processing was developed and evaluated. Whole blood was collected on filter paper that lysed cells and bound the DNA, eliminating specimen centrifugation and extraction procedures. The DNA remained bound to the filter paper during PCR amplification. Assays of copy number standards showed reproducible detection of 5 to 10 copies of HIV-1 in 5 microl of whole blood. The sensitivity of the assay did not decrease after storage of the standards on filter paper for 3 months at room temperature or after incubation at 37 or 45 degrees C for 20 h. The primers used for nested PCR of the HIV-1 pol gene amplified templates from a reference panel of multiple HIV-1 subtypes but did not amplify a subtype A or a subtype C virus from children living in Seattle. The assay had a sensitivity of 98.4% and a specificity of 98.3% for testing of 122 specimens from 35 HIV-1-infected and 16 uninfected children and 43 seronegative adults living in Washington. The assay had a sensitivity of 99% and a specificity of 100% for testing of 102 HIV-1-positive (as determined by enzyme immunoassay) Peruvian women and 6 seropositive and 34 seronegative infants. This assay, with adsorption of whole blood to filter paper and no specimen processing, provides a practical, economical, sensitive, and specific method for the diagnosis of HIV-1 subtype B infection in infants.

Oral ganciclovir in children: pharmacokinetics, safety, tolerance, and antiviral effects. The Pediatric AIDS Clinical Trials Group.

The pharmacokinetics, safety, tolerance, and antiviral effects of ganciclovir (Gcv) administered orally were evaluated in 36 children infected with cytomegalovirus (CMV) who were severely immunocompromised by infection with human immunodeficiency virus type 1. In this dose-escalation study, 30 mg/kg of Gcv administered every 8 h produced serum levels similar to the dose (1 g/8 h) effective for maintenance treatment of CMV retinitis in adults. In older children, serum Gcv concentrations were similar after the administration of capsules and suspension. All doses (10-50 mg/kg/8 h) studied were safe and, except for the volume of suspension or number of pills, were well tolerated. Oral Gcv was associated with a decrease in the detection of CMV by culture or polymerase chain reaction. CMV disease occurred in 3 children during the study: one developed Gcv resistance, another had harbored resistant virus at study entry, and a third had wild-type CMV

Pediatric human immunodeficiency virus type 1 infection: updates on prevention and management.

A greater understanding of perinatal HIV-1 transmission has resulted in a marked decrease in the incidence of pediatric HIV-1 infection in several affluent nations. Advances in treatment have led to a decrease in HIV-1-related morbidity and mortality in some HIV-1-infected children. Studies are ongoing to determine methods to sustain the salutary effects of treatment. A goal of child advocates is to translate these advances to communities with few resources. Efforts are underway to make infant chemoprophylaxis widely available and to define strategies to reduce exposure to maternal virus. Likewise, there is the need to increase the access of children to treatment.

Anti-retroviral therapy for HIV-infected children--toward maximal effectiveness.

Viral suppression may be more difficult to achieve in children compared to adults. However, appropriate regimens at adequate dosages combined with monitoring of adherence and virologic response should allow children with HIV infection to realize a sustained benefit from combination anti-retroviral therapy.

HIV-1 dynamics in children.

HIV-1-infected children have higher plasma viral loads and progress to disease more quickly than infected adults. To gain insight into the accelerated pathogenesis of HIV-1 in children, viral dynamics were measured following the initiation of highly active antiretroviral therapy (HAART) and compared with those reported for adults. A biphasic decline in plasma HIV-1 RNA was observed, with a rapid decrease during the first 1 to 2 weeks of therapy (phase I) followed by a slower decline (phase II). The phase I and II decay rates were not significantly different among children of different ages, pretherapy plasma HIV-1 RNA levels, or CD4 cell counts. Estimated phase I decay rates were similar to those previously reported in adults with a mean of 0.43 days(-1) and a half-life of 1.6 days. The phase II decay rates were slower in children compared with adults with a mean of 0.016 days(-1) versus 0.066 days(-1), and a half-life of 43.3 versus 14.1 days, respectively (p < .05). The mean time required to reach viral levels below detection thresholds was also longer in these children compared with that in adults. These data suggest that HIV-1 dynamics may be different in children, and that these differences may necessitate different treatment strategies.

Risk of mother-to-infant transmission of HIV-1 is not reduced in CCR5/delta32ccr5 heterozygotes.

To determine if the 32-bp deletion of the chemokine receptor CCR5 (delta32ccr5) protects against mother-to-infant transmission of HIV-1, specimens from all uninfected and infected children who were perinatally exposed to HIV-1 and observed since 1988 and whose mothers did not take zidovudine were assessed for delta32ccr5. The CCR5 genotype was determined using polymerase chain reaction (PCR) for 122 subjects, of whom 73 were HIV-1 infected and 49 were perinatally exposed but uninfected; 70% and 71%, respectively, were Caucasian. Eleven of 73 (15%) infected children and 4 of 49 (8%) exposed uninfected children were CCR5/delta32ccr5 heterozygotes (p = 0.40). Among subjects who had at least one Caucasian parent or grandparent, 11 of 51 (22%) HIV-1-infected persons and 4 of 35 (11%) uninfected persons were heterozygotes. None were homozygous for the delta32ccr5 allele. The estimated relative risk for mother-to-infant HIV-1 transmission in heterozygotes was 2.0. Furthermore, the 95% confidence interval (0.6, 7.3) suggested that it is unlikely that the true relative risk was <0.6. Thus, the infant CCR5/delta32ccr5 heterozygous genotype was not associated with a diminished risk of perinatally acquired HIV-1 infection.

Clinical, virologic and immunologic responses of children with advanced human immunodeficiency virus type 1 disease treated with protease inhibitors.

OBJECTIVE: To determine the effects of combination antiretroviral therapy including a protease inhibitor (PI combination therapy) in children with advanced HIV-1 disease. STUDY DESIGN: An observational study of HIV-1 plasma RNA, lymphocyte subsets, delayed type hypersensitivity and physical growth after initiation of PI combination therapy. RESULTS: In nine children the median HIV-1 plasma RNA decreased 1.7 log10 (mean, 1.57; range, 0.7 to 2.2) following PI combination therapy and CD4 cells increased a median of 499 (mean, 528; range, 9 to 1088) cells/microl. A rebound of RNA, associated with the development of resistance to the PI, occurred in three subjects. Three of six children were no longer anergic and all nine achieved normal weight-growth velocities. Ritonavir was well-tolerated, despite its bitter taste; however, four of five children treated with indinavir developed renal complications. CONCLUSIONS: PI combination therapy in children with advanced HIV-1 disease was associated with a decrease in HIV-1 RNA, improved immunologic measures and normal or better weight gain. Of concern was the rebound in plasma HIV-1 associated with resistance to the PI observed in one-third of patients. This emphasizes the need for larger studies to define optimal PI containing regimens with long term efficacy in children.

Prevention of mother-to-infant transmission of HIV-1.

The number of children with AIDS continues to increase worldwide. Children who become infected with HIV-1 acquire the infection almost exclusively from their mothers during pregnancy or delivery, or via breast feeding. Mother-to-infant transmission has been, and continues to be, an area of active research with the goal being complete prevention. Treatment with zidovudine, an antiviral agent, has been found to decrease transmission from 25% to 8%. However, multiple obstacles impede the worldwide application of this advance.

Analysis of the maternal components of the AIDS clinical trial group 076 zidovudine regimen in the prevention of mother-to-infant transmission of human immunodeficiency virus type 1.

To gain insight into the protective effects of the three components of the zidovudine regimen used in AIDS Clinical Trial Group (ACTG) 076 on mother-to-infant transmission of human immunodeficiency virus (HIV) type 1, 188 zidovudine-treated women and their untreated infants from five HIV-1 obstetric centers were retrospectively studied. The overall rate of mother-to-infant transmission was 12.3% (95% confidence interval [CI], 7.9%-18.0%). When the 38 women with <200 CD4 cells/microL were excluded, the mother-to-infant transmission rate was 8.8% (95% CI, 4.6%-14.8%). This rate compares favorably with the 8.3% transmission in the zidovudine arm of the ACTG 076 study. Apart from low (<200/microL) maternal CD4 cells (P = .016), no factors, including the duration of zidovudine therapy during gestation and intravenous administration of zidovudine during labor, affected the rate of mother-to-infant transmission. These findings suggest that antenatal oral zidovudine may be as effective as antenatal oral plus intravenous zidovudine during labor and the three-component ACTG 076 regimen in decreasing mother-to-infant HIV-1 transmission.

Lack of detection of human immunodeficiency virus type 1 in the saliva of infected children and adolescents.

OBJECTIVE: To determine the prevalence of human immunodeficiency virus type 1 (HIV-1) in the saliva of infected children and adolescents. METHODS: Saliva and blood samples were collected from 13 patients (age range, 1-15 years) with HIV-1 infection. Eleven were taking antiretroviral agents. The presence of HIV-1 was determined by polymerase chain reaction analysis of RNA and DNA as well as by viral culture of the saliva samples and by culture of peripheral blood mononuclear cells. RESULTS: Although HIV-1 was cultured from peripheral blood mononuclear cells of 12 patients, it was not cultured from their saliva. Only 1 of 13 saliva samples yielded positive test results for HIV-1 RNA, and none did so for HIV-1 DNA. The specimen containing HIV-1 RNA was from an untreated 10-year-old asymptomatic boy with a CD4+ lymphocyte count of 0.91 x 10(9)/L (913 cells/microL) and no infectious virus detected in plasma. CONCLUSION: The prevalence of HIV-1 in the saliva of HIV-1-infected children and adolescents is low and may not be infectious.

Effect of pregnancy and zidovudine therapy on viral load in HIV-1-infected women.

The objective of this study was to determine the effect of pregnancy and zidovudine (ZDV) on viral load in HIV-1 infected women. A prospective nonrandomized cohort study was conducted at a university medical center and affiliated clinic and included 44 HIV-1-seropositive pregnant women seen between June 1991 and September 1995. Twenty-three women initiated ZDV therapy during their pregnancy. Seventeen women did not take antiretrovirals, and four women took ZDV prior to and throughout pregnancy. HIV-1 viral load as determined by quantitative peripheral blood mononuclear cell (PBMC) culture and quantitative plasma RNA levels was measured at various times during pregnancy and in the postpartum period. HIV-1 load, by both infectivity and RNA levels, was relatively low and remained stable during pregnancy and through 6 weeks post partum. Initiation of ZDV therapy during pregnancy did not result in a significant decrease in viral load at delivery when controlling for the effect of pregnancy. In those women who received ZDV therapy only during pregnancy, there was a trend toward an increase in viral load measured by PBMC infectivity 6 months post partum compared with the levels before the initiation of ZDV. Mother-to-child transmission of HIV-1 occurred in one of 27 (4%) ZDV-treated women and in two of 16 (12.5%) untreated women. Among HIV-1-infected pregnant women with low viral levels, HIV-1 plasma RNA and infectivity remained stable during and after gestation. Although these results are based on a relatively small number of women and should be considered preliminary, the lack of significant ZDV-associated diminution in viral levels suggests that the protective effect of ZDV on the mother-to-child transmission of HIV-1 may not be due to the reduction in maternal viral levels but, by inference, may be due to the prevention of HIV-1 reverse transcription in the newborn.

Pediatric HIV disease: new developments in treatment and prevention.

Despite the continued increase in the number of HIV-1-infected children, there is optimism that with the prospective identification of HIV-1-infected pregnant women, the incidence of infection can be diminished through prevention of mother-to-child transmission both by use of antiretroviral agents as well as by improved obstetrical management. Also, insight into viral dynamics and the advent of more potent antiretroviral agents provides hope for better treatment strategies and thus the ability to slow disease progression of HIV-1-infected children.

Pediatric HIV-1 infection.

Although there is optimism that with the prospective identification and treatment of HIV-1-infected pregnant women the incidence of pediatric infection can be diminished, currently the number of HIV-1-infected children continues to rise. Improvements in early diagnosis provide the potential for early intervention, and the advent of more potent antiretroviral agents provides the hope of better treatment strategies to slow disease progression in HIV-1-infected children.

Hypomethylation of the interferon-gamma gene correlates with its expression by primary T-lineage cells.

To determine the potential role of methylation in the regulation of interferon-gamma (IFN-gamma) gene transcription by T cells, primary T-lineage cell populations were analyzed for the extent of methylation of three CpG sites within or near transcriptional activator elements in the 5' flank and first intron of the human IFN-gamma gene. A striking correlation was observed between the capacity of the IFN-gamma gene to be expressed and the degree of hypomethylation. The IFN-gamma gene was virtually completely methylated at all sites in thymocytes, neonatal T cells, and adult CD45RAhiCD45R0lo (antigenically naive) CD4 T cells, cell types that all have a low or undetectable capacity to express the IFN-gamma gene. In contrast, there was substantial hypomethylation in T-lineage cell types with relatively high capacities to express the IFN-gamma gene, including adult CD8 T cells and adult CD45RAloCD45R0hi (memory/effector) CD4 T cells. These results suggest that hypomethylation of the IFN-gamma genetic locus may be an important determinant of IFN-gamma gene expression in vivo by T-lineage cells.

Differentiation of the T helper phenotypes by analysis of the methylation state of the IFN-gamma gene.

Th1 and Th2 CD4+ T cell clones have been defined by their ability to produce different lymphokines. However, the processes by which CD4+ T cells differentially regulate lymphokine gene expression have not been well defined. In this report, we demonstrate that the methylation status of a CpG dinucleotide contained within a TATA proximal regulatory element of the IFN-gamma promoter correlates with the transcription of the gene. In murine Th1 clones and two human CD4+ Th0 clones, this site is either completely or partially hypomethylated, whereas in murine Th2 clones this site is > 98% methylated. Treatment of murine Th2 clones with 5-azacytidine, an agent that inhibits methylation of the DNA, converts these cells to IFN-gamma producers. Additional targets for methylation outside the transcriptional control regions of the IFN-gamma genetic locus were found to be hypomethylated in Th2 cells but not in Th1 cells. Electrophoretic mobility shift assays (EMSA) revealed at least five distinct protein-DNA complexes that are formed with an oligonucleotide containing the IFN-gamma promoter TATA proximal regulatory element, and in vitro methylation of this site results in a loss of these three complexes. Furthermore, a comparison of nuclear extracts prepared from Th1 and Th2 clones revealed that the EMSA patterns were qualitatively similar but differed quantitatively. In addition, transient transfection of a murine IFN-gamma promoter-chloramphenicol acetyl transferase (CAT) gene construct into both Th1 and Th2 clones produced CAT activity that was not inducible by anti-CD3, indicating that hypomethylation per se of the promoter alone is not sufficient for inducible gene expression.