OBJECTIVE

To identify the effects of substance abuse status (active, former, and never) on utilization of highly active antiretroviral therapy (HAART), medication adherence, and virologic and immunologic responses to therapy.

METHODS

Prospective cohort study of 764 HIV-<em>1</em>-infected patients who attended an urban HIV clinic and participated in a standardized interview.

METHODS

Past utilization of HAART, self-reported nonadherence with antiretroviral therapy, and changes in HIV-<em>1</em> RNA level and CD4+ lymphocyte count relative to prior peak and nadir, respectively.

RESULTS

Forty-four percent of active drug users failed to utilize HAART compared with 22% of former drug users and <em>1</em>8% of non-drug users (p <.00<em>1</em> for both comparisons). Among participants who were taking antiretroviral therapy when interviewed, active drug users were more likely to report medication nonadherence (34% vs. 24% of nonusers and <em>1</em>7% of former users), had a smaller median reduction in HIV-<em>1</em> RNA from baseline (0.8 log<em>1</em>0 copies/ml vs. <em>1</em>.7 in nonusers and <em>1</em>.6 in former users), and had smaller median increases in CD4+ lymphocyte count from baseline (65 cells/mm3 vs. <em>1</em><em>1</em>6 in nonusers and <em>1</em>22 in former users) (p <.05 for all comparisons with active users).

CONCLUSIONS

Active drug use was strongly associated with underutilization of HAART, nonadherence, and inferior virologic and immunologic responses to therapy, whereas former drug users and non-drug users were similar in all outcomes. Effective strategies are needed that integrate HIV-<em>1</em> and substance abuse treatments.

In vitro studies show that human immunodeficiency virus type <em>1</em> (HIV-<em>1</em>) does not replicate in freshly isolated monocytes unless monocytes differentiate to monocyte-derived macrophages. Similarly, HIV-<em>1</em> may replicate in macrophages in vivo, whereas it is unclear whether blood monocytes are permissive to productive infection with HIV-<em>1</em>. We investigated HIV-<em>1</em> replication in CD<em>1</em>4(+) monocytes and resting and activated CD4(+) T cells by measuring the levels of cell-associated viral DNA and mRNA and the genetic evolution of HIV-<em>1</em> in seven acutely infected patients whose plasma viremia had been (<em>1</em>00 copies/<em>ml</em> for 803 to <em>1</em>,544 days during highly active antiretroviral therapy (HAART). HIV-<em>1</em> DNA was detected in CD<em>1</em>4(+) monocytes as well as in activated and resting CD4(+) T cells throughout the course of study. While significant variation in the decay slopes of HIV-<em>1</em> DNA was seen among individual patients, viral decay in CD<em>1</em>4(+) monocytes was on average slower than that in activated and resting CD4(+) T cells. Measurements of HIV-<em>1</em> sequence evolution and the concentrations of unspliced and multiply spliced mRNA provided evidence of ongoing HIV-<em>1</em> replication, more pronounced in CD<em>1</em>4(+) monocytes than in resting CD4(+) T cells. Phylogenetic analyses of HIV-<em>1</em> sequences indicated that after prolonged HAART, viral populations related or identical to those found only in CD<em>1</em>4(+) monocytes were seen in plasma from three of the seven patients. In the other four patients, HIV-<em>1</em> sequences in plasma and the three cell populations were identical. CD<em>1</em>4(+) monocytes appear to be one of the potential in vivo sources of HIV-<em>1</em> in patients receiving HAART.

To examine the role of adipose-resident macrophages in insulin resistance, we examined the gene expression of CD68, a macrophage marker, along with macrophage chemoattractant protein-<em>1</em> (MCP-<em>1</em>) in human subcutaneous adipose tissue using real-time RT-PCR. Both CD68 and MCP-<em>1</em> mRNAs were expressed in human adipose tissue, primarily in the stromal vascular fraction. When measured in the adipose tissue from subjects with normal glucose tolerance, covering a wide range of BMI (2<em>1</em>-5<em>1</em> kg/m2) and insulin sensitivity (S(I)) (0.6-8.0 x <em>1</em>0(-4)min(-<em>1</em>).microU(-<em>1</em>).<em>ml</em>(-<em>1</em>)), CD68 mRNA abundance, which correlated with the number of CD68-positive cells by immunohistochemistry, tended to increase with BMI but was not statistically significant. However, there was a significant inverse relation between CD68 mRNA and S(I) (r=-0.55, P=0.02). In addition, there was a strong positive relationship among adipose tissue CD68 mRNA, tumor necrosis factor-alpha (TNF-alpha) secretion in vitro (r=0.79, P<0.005), and plasma interleukin-6 (r=0.67, P < 0.005). To determine whether improving S(I) in subjects with impaired glucose tolerance (IGT) was associated with decreased CD68 expression, IGT subjects were treated for <em>1</em>0 weeks with pioglitazone or metformin. Pioglitazone increased S(I) by 60% and in the same subjects reduced both CD68 and MCP-<em>1</em> mRNAs by >50%. Furthermore, pioglitazone resulted in a reduction in the number of CD68-positive cells in adipose tissue and reduced plasma TNF-alpha. Metformin had no effect on any of these measures. Thus, treatment with pioglitazone reduces expression of CD68 and MCP-<em>1</em> in adipose tissue, apparently by reducing macrophage numbers, resulting in reduced inflammatory cytokine production and improvement in S(I).

OBJECTIVE

We assessed the feasibility of noninvasive metabolic monitoring of cancer chemohormonotherapy using sequential quantitative positron emission tomographic (PET) scans of tumor glucose metabolism with the glucose analog 2-[<em>1</em>8F]-fluoro-2-deoxy-D-glucose (FDG).

METHODS

Eleven women with newly diagnosed primary breast cancers larger than 3 cm in diameter beginning a chemohormonotherapy program underwent a baseline and four follow-up quantitative PET scans during the first three cycles of treatment (days 0 to 63). Tumor response was sequentially determined clinically, radiographically, and then pathologically after nine treatment cycles.

RESULTS

Eight patients had partial or complete pathologic responses. Their maximal tumor uptake of FDG assessed by PET decreased promptly with treatment to the following: day 8, 78 +/- 9.2% (P < .03); day 2<em>1</em>, 68.<em>1</em> +/- 7.5% (P < .025); day 42, 60 +/- 5.<em>1</em>% (P < .00<em>1</em>); day 63, 52.4 +/- 4.4% (P < .000<em>1</em>) of the basal values. Tumor diameter did not decrease significantly during this period through 63 days. Prompt decreases in the FDG influx rate (K) from basal levels (from .0<em>1</em>9 to .0<em>1</em>4 <em>mL</em>/cm3/min) after 8 days of treatment (P < .02) and in the estimated rate of FDG phosphorylation to FDG-6-phosphate (k3) from .055 to .038 min-<em>1</em> after 8 days of treatment (P < .02) to .029 +/- .004 min-<em>1</em> at 2<em>1</em> days) (P < .02) were observed. Three nonresponding patients had no significant decrease in tumor uptake of FDG (8<em>1</em> +/- <em>1</em>8% of basal value), influx rate (.0<em>1</em>5 to .0<em>1</em>2 <em>mL</em>/cm3/min), or tumor size (8<em>1</em> +/- <em>1</em>2% of basal diameter) comparing basal versus 63-day posttreatment values.

CONCLUSIONS

Quantitative FDG PET scans of primary breast cancers showed a rapid and significant decrease in tumor glucose metabolism after effective treatment was initiated, with the reduction in metabolism antedating any decrement in tumor size. No significant decrease in FDG uptake (SUV) after three cycles of treatment was observed in the nonresponding patients. FDG PET scanning has substantial promise as an early noninvasive metabolic marker of the efficacy of cancer treatment.

A circulating form of the usually membrane-bound intercellular adhesion molecule-<em>1</em> (ICAM-<em>1</em>) was identified and characterized in normal human serum, and in sera from patients with leukocyte adhesion deficiency (LAD). The molecule, designated circulating ICAM-<em>1</em> (cICAM-<em>1</em>) was detected and quantitated by sandwich ELISA. Levels of cICAM-<em>1</em> in sera from normal individuals ranged from <em>1</em>00 to 200 ng/<em>ml</em>. Sera from LAD patients had elevated cICAM-<em>1</em> levels ranging from 200 to 700 ng/<em>ml</em>. The elevated levels of cICAM-<em>1</em> in LAD sera may be due to an inability to adsorb cICAM-<em>1</em> to cell-bound LFA-<em>1</em> or may be an indirect result of the pathology accompanying the syndrome. cICAM-<em>1</em> bound to mAb specific for four distinct ICAM-<em>1</em> epitopes localized in domains D<em>1</em>, D2, D4, and D5, and displayed similar molecular size properties as recombinant soluble ICAM-<em>1</em> on FPLC size-exclusion chromatography. When immobilized via a domain D5-specific mAb, cICAM-<em>1</em> mediated function (LFA-<em>1</em>)-dependent lymphocyte adhesion equivalent to sICAM-<em>1</em>. These data indicate that cICAM-<em>1</em> contains most, if not all, of the five extracellular domains of membrane ICAM-<em>1</em>, as well as the ability to bind specifically to LFA-<em>1</em>. The cellular source of cICAM-<em>1</em> appeared to be from mononuclear cells; only lymphoid cell lines or primary PBMC cultures had detectable levels of cICAM-<em>1</em> in cell culture supernatants. Because cICAM-<em>1</em> retains the ability to bind specifically to LFA-<em>1</em>, it may act to regulate cell adhesion by promoting de-adhesion. Alternatively, cICAM-<em>1</em> may be the indirect consequence of inflammation or tissue damage. As such, the detection of cICAM-<em>1</em> could be useful as a marker of inflammatory disease.

Measurements of the attachment of the PAM 2<em>1</em>2 line of mouse epithelial cells to various collagen substrates show that these cells adhere preferentially to type IV, basement membrane collagen. Neither serum nor fibronectin stimulated the attachment of these cells (unlike fibroblasts) to type IV collagen. Preincubation of the PAM 2<em>1</em>2 cells with cycloheximide prevented attachment. Thus these cells do not attach by means of a macromolecule present inserum, but instead synthesize an attachment factor. Extracts of the EHS tumor, which produces an extracellular matrix containing basement membrane components, were tested for their ability to promote attachment to cycloheximide-treated PAM 2<em>1</em>2 cells. Saline extracts of the tumor stimulated the attachment of the PAM 2<em>1</em>2 cells to type IV collagen in the presence of cycloheximide. Laminin, a high molecular weight glycoprotein constituent of basement membrane, was purified from the salt extract and was found to be the active species at concentrations as low as <em>1</em>-5 microgram/<em>ml</em>. When laminin was preincubated on plates coated with either type I, II, III, IV, or V collagen and the plates subsequently washed, high levels of attachment were seen only on type IV collagen-coated plates. Affinity purified antibody directed against laminin inhibited the attachment of PAM 2<em>1</em>2 cells to a type IV collagen substrate. Laminin appears to be a specific attachment protein for epithelial cells since it did not stimulate the attachment of fibroblasts to type I or to type IV collagen substrates. These data suggest that lamin is produced and utilized by these epithelial cells to attach to basement membrane collagen.

OBJECTIVE

To evaluate survival and investigate causes of death among HIV-<em>1</em> infected adults receiving HAART in Senegal.

METHODS

An observational prospective cohort.

METHODS

Mortality was assessed in the first patients enrolled between August <em>1</em>998 and April 2002 in the Senegalese antiretroviral drug access initiative. First-line regimen combined two nucleoside reverse transcriptase inhibitors and either a non-nucleoside reverse transcriptase inhibitor or a protease inhibitor. The most likely causes of death were ascertained through medical records or post-mortem interviews (verbal autopsy).

RESULTS

Four hundred and four patients (54.7% women) were enrolled in the study and were followed for a median of 46 months (interquartile range: 32-57 months) after HAART initiation. At baseline, 5% were antiretroviral therapy (ART) non-naive, 39 and 55% were respectively at CDC stage B and C, median age, CD4 cell count and viral load were 37 years, <em>1</em>28 cells/microl and 5.2 log cp/ml, respectively. Ninety-three patients died during follow-up and the overall incidence rate of death was 6.3/<em>1</em>00 person-years [95% confidence interval (CI), 5.2-7.7]. During the first year after HAART initiation, 47 patients died and seven were lost to follow-up, yielding to a probability of dying of <em>1</em><em>1</em>.7% (95% CI, 8.9-<em>1</em>5.3%). The death rate, which was highest during the first year after HAART initiation, decreased with time yielding a cumulative probability of dying of <em>1</em>7.4% (95% CI, <em>1</em>3.9-2<em>1</em>.5%) and 24.6% (95% CI, 20.4-29.4%) at 2 and 5 years. Causes of death were ascertained in 76 deaths. Mycobacterial infections, neurotropic infections and septicaemia were the most frequent likely causes of death.

CONCLUSIONS

This study underlines the early mortality pattern after HAART initiation and highlights the leading role of mycobacterial infections in the causes of death.

We have previously demonstrated that the Gag p9 protein of equine infectious anemia virus (EIAV) is functionally homologous with Rous sarcoma virus (RSV) p2b and human immunodeficiency virus type <em>1</em> (HIV-<em>1</em>) p6 in providing a critical late assembly function in RSV Gag-mediated budding from transfected COS-<em>1</em> cells (L. J. Parent et al., J. Virol. 69:5455-5460, <em>1</em>995). In light of the absence of amino acid sequence homology between EIAV p9 and the functional homologs of RSV and HIV-<em>1</em>, we have now designed an EIAV Gag-mediated budding assay to define the late assembly (L) domain peptide sequences contained in the EIAV p9 protein. The results of these particle budding assays revealed that expression of EIAV Gag polyprotein in COS-<em>1</em> cells yielded extracellular Gag particles with a characteristic density of <em>1</em>.<em>1</em>8 g/<em>ml</em>, while expression of EIAV Gag polyprotein lacking p9 resulted in a severe reduction in the release of extracellular Gag particles. The defect in EIAV Gag polyprotein particle assembly could be corrected by substituting either the RSV p2b or HIV-<em>1</em> p6 protein for EIAV p9. These observations demonstrated that the L domains of EIAV, HIV-<em>1</em>, and RSV were interchangeable in mediating assembly of EIAV Gag particles in the COS-<em>1</em> cell budding assay. To localize the L domain of EIAV p9, we next assayed the effects of deletions and site-specific mutations in the p9 protein on its ability to mediate budding of EIAV Gag particles. Analyses of EIAV Gag constructs with progressive N-terminal or C-terminal deletions of the p9 protein identified a minimum sequence of <em>1</em><em>1</em> amino acids (Q20N2<em>1</em>L22Y23P24D25L26S27E28I29K30) capable of providing the late assembly function. Alanine scanning studies of this L-domain sequence demonstrated that mutations of residues Y23, P24, and L26 abrogated the p9 late budding function; mutations of other residues in the p9 L domain did not substantially affect the level of EIAV Gag particle assembly. These data indicate that the L domain in EIAV p9 utilizes a YXXL motif which we hypothesize may interact with cellular proteins to facilitate virus particle budding from infected cells.

BACKGROUND

Human papillomavirus (HPV) infection is associated with precancerous cervical squamous intraepithelial lesions commonly seen among women infected with human immunodeficiency virus-<em>1</em> (HIV). We characterized HPV infection in a large cohort of HIV-positive and HIV-negative women participating in the Women's Interagency HIV Study to determine the prevalence of and risk factors for cervicovaginal HPV infection in HIV-positive women.

METHODS

HIV-positive (n = <em>1</em>778) and HIV-negative (n = 500) women were tested at enrollment for the presence of HPV DNA in a cervicovaginal lavage specimen. Blood samples were tested for HIV antibody status, level of CD4-positive T cells, and HIV RNA load (copies/mL). An interview detailing risk factors was conducted. Univariate and multivariate analyses were performed.

RESULTS

Compared with HIV-negative women, HIV-positive women with a CD4+ cell count of less than 200/mm3 were at the highest risk of HPV infection, regardless of HIV RNA load (odds ratio [OR] = <em>1</em>0.<em>1</em>3; 95% confidence interval [CI] = 7.32-<em>1</em>4.04), followed by women with a CD4+ count greater than 200/mm3 and an HIV RNA load greater than 20,000 copies/mL (OR = 5.78; 95% CI = 4.<em>1</em>7-8.08) and women with a CD4+ count greater than 200/mm3 and an HIV RNA load less than 20,000 copies/mL (OR = 3.<em>1</em>2; 95% CI = 2.36-4.<em>1</em>2), after adjustment for other factors. Other risk factors among HIV-positive women included racial/ethnic background (African-American versus Caucasian, OR = <em>1</em>.64; 95% CI = <em>1</em>.<em>1</em>9-2.28), current smoking (yes versus no; OR = <em>1</em>.55; 95% CI = <em>1</em>.20-<em>1</em>.99), and younger age (age < 30 years versus>> or = 40 years; OR = <em>1</em>.75; 95% CI = <em>1</em>.23-2.49).

CONCLUSIONS

Although the strongest risk factors of HPV infection among HIV-positive women were indicators of more advanced HIV-related disease, other factors commonly found in studies of HIV-negative women, including racial/ethnic background, current smoking, and age, were important in HIV-positive women as well.

The recent isolation of the ammonia-oxidizing crenarchaeon Nitrosopumilus maritimus has expanded the known phylogenetic distribution of nitrifying phenotypes beyond the domain Bacteria. To further characterize nitrification in the marine environment and explore the potential crenarchaeal contribution to this process, we quantified putative nitrifying genes and phylotypes in picoplankton genomic libraries and environmental DNA samples from coastal and open ocean habitats. Betaproteobacteria ammonia monooxygenase subunit A (amoA) gene copy numbers were low or undetectable, in stark contrast to crenarchaeal amoA-like genes that were broadly distributed and reached up to 6 x <em>1</em>0(4) copies <em>ml</em>(-<em>1</em>). Unexpectedly, in the North Pacific Subtropical Gyre, a deeply branching crenarchaeal group related to a hot spring clade (pSL<em>1</em>2) was at times abundant below the euphotic zone. Quantitative data suggested that the pSL<em>1</em>2 relatives also contain archaeal amoA-like genes. In both coastal and open ocean habitats, close relatives of known nitrite-oxidizing Nitrospina species were well represented in genomic DNA libraries and quantitative PCR profiles. Planktonic Nitrospina depth distributions correlated with those of Crenarchaea. Overall, the data suggest that amoA-containing Crenarchaea are more phylogenetically diverse than previously reported. Additionally, distributional patterns of planktonic Crenarchaea and Nitrospina species suggest potential metabolic interactions between these groups in the ocean's water column.

The incidence of ESRD is increasing dramatically. Progression to end-stage may be halted or slowed when kidney damage is detected at an early stage. Kidney damage is frequently asymptomatic but is indicated by the presence of proteinuria, hematuria, or reduced GFR. Population-based studies relating to the prevalence of kidney damage in the community are limited, particularly outside of the United States. Therefore, the prevalence of proteinuria, hematuria, and reduced GFR in the Australian adult population was determined using a cross-sectional study of <em>1</em><em>1</em>,247 noninstitutionalized Australians aged 25 yr or over, rando<em>ml</em>y selected using a stratified, cluster method. Subjects were interviewed and tested for proteinuria-spot urine protein to creatinine ratio (abnormal:>>/=0.20 mg/mg); hematuria-spot urine dipstick (abnormal: <em>1</em>+ or greater) confirmed by urine microscopy (abnormal:>><em>1</em>0,000 red blood cells per milliliter) or dipstick (abnormal: <em>1</em>+ or greater) on midstream urine sample; and reduced GFR-Cockcroft-Gault estimated GFR (abnormal: <60 <em>ml</em>/min per <em>1</em>.73 m(2)). The associations between age, gender, diabetes mellitus, and hypertension, and indicators of kidney damage were examined. Proteinuria was detected in 2.4% of cases (95% CI: <em>1</em>.6%, 3.<em>1</em>%), hematuria in 4.6% (95% CI: 3.8%, 5.4%), and reduced GFR in <em>1</em><em>1</em>.2% (95% CI: 8.6%, <em>1</em>3.8%). Approximately <em>1</em>6% had at least one indicator of kidney damage. Age, diabetes mellitus, and hypertension were independently associated with proteinuria; age, gender, and hypertension with hematuria; and age, gender, and hypertension with reduced GFR. Approximately <em>1</em>6% of the Australian adult population has either proteinuria, hematuria, and/or reduced GFR, indicating the presence of kidney damage. Identifying and targeting this section of the population may provide a means to reduce the burden of ESRD.

Plasma leptin concentration is increased in hypertensive obese humans, but whether leptin contributes to the increased arterial pressure in obesity is not known. In this study, we tested whether chronic increases in leptin, to levels comparable to those in obesity, could cause a sustained increase in arterial pressure and also the importance of central nervous system (CNS) versus systemic mechanisms. Five male Sprague-Dawley rats were implanted with chronic nonoccluding catheters in the abdominal aorta and both carotid arteries for CNS infusion, and five other rats were implanted with an abdominal aorta catheter and femoral vein catheter for intravenous (I.V.) infusion. After 7 days of control, leptin was infused into the carotid arteries or femoral vein at 0.<em>1</em> microg/kg/min for 5 days and <em>1</em>.0 microg/kg/min for 7 days, followed by a 7-day recovery period. The carotid artery and i.v. infusions of leptin at <em>1</em> microg/kg/min significantly increased plasma leptin levels, from <em>1</em>.2+/-0.4 ng/<em>mL</em> to 9<em>1</em>+/-5 ng/<em>mL</em> and from 0.9+/-0.<em>1</em> ng/<em>mL</em> to 94+/-9 ng/<em>mL</em>, respectively, but there was no significant increase in either group at the low dose. Food intake also did not change at the low dose but decreased by approximately 65% in the carotid group and 69% in the i.v. group after 7 days of the <em>1</em> microg/kg/min infusion. Mean arterial pressure (MAP) increased slightly at the low dose only in the carotid group, but this was not statistically significant. At the higher dose, however, MAP increased significantly from 86+/-<em>1</em> mm Hg to 94+/-<em>1</em> mm Hg in the carotid group and from 87+/-<em>1</em> mm Hg to 93+/-<em>1</em> mm Hg in the i.v. group. Heart rate also increased significantly in both groups at <em>1</em> microg/kg/min leptin infusion. Fasting blood glucose and insulin levels decreased significantly at <em>1</em> microg/kg/min in both the carotid artery group (-<em>1</em>0.5% and -82.5%, respectively) and the i.v. group (-<em>1</em>3.6% and -80.4%, respectively). All variables returned to control levels after leptin infusion was stopped. These results indicate that chronic increases in circulating leptin cause sustained increases in arterial pressure and heart rate and are consistent with a possible role for leptin in obesity hypertension.

OBJECTIVE

A key factor in assessing the effectiveness and cost-effectiveness of antiretroviral therapy (ART) as a prevention strategy is the absolute risk of HIV transmission through condomless sex with suppressed HIV-<em>1</em> RNA viral load for both anal and vaginal sex.

OBJECTIVE

To evaluate the rate of within-couple HIV transmission (heterosexual and men who have sex with men [MSM]) during periods of sex without condoms and when the HIV-positive partner had HIV-<em>1</em> RNA load less than 200 copies/mL.

METHODS

The prospective, observational PARTNER (Partners of People on ART-A New Evaluation of the Risks) study was conducted at 75 clinical sites in <em>1</em>4 European countries and enrolled <em>1</em><em>1</em>66 HIV serodifferent couples (HIV-positive partner taking suppressive ART) who reported condomless sex (September 20<em>1</em>0 to May 20<em>1</em>4). Eligibility criteria for inclusion of couple-years of follow-up were condomless sex and HIV-<em>1</em> RNA load less than 200 copies/mL. Anonymized phylogenetic analysis compared couples' HIV-<em>1</em> polymerase and envelope sequences if an HIV-negative partner became infected to determine phylogenetically linked transmissions.

METHODS

Condomless sexual activity with an HIV-positive partner taking virally suppressive ART.

METHODS

Risk of within-couple HIV transmission to the HIV-negative partner.

RESULTS

Among <em>1</em><em>1</em>66 enrolled couples, 888 (mean age, 42 years [IQR, 35-48]; 548 heterosexual [6<em>1</em>.7%] and 340 MSM [38.3%]) provided <em>1</em>238 eligible couple-years of follow-up (median follow-up, <em>1</em>.3 years [IQR, 0.8-2.0]). At baseline, couples reported condomless sex for a median of 2 years (IQR, 0.5-6.3). Condomless sex with other partners was reported by <em>1</em>08 HIV-negative MSM (33%) and 2<em>1</em> heterosexuals (4%). During follow-up, couples reported condomless sex a median of 37 times per year (IQR, <em>1</em>5-7<em>1</em>), with MSM couples reporting approximately 22,000 condomless sex acts and heterosexuals approximately 36,000. Although <em>1</em><em>1</em> HIV-negative partners became HIV-positive (<em>1</em>0 MSM; <em>1</em> heterosexual; 8 reported condomless sex with other partners), no phylogenetically linked transmissions occurred over eligible couple-years of follow-up, giving a rate of within-couple HIV transmission of zero, with an upper 95% confidence limit of 0.30/<em>1</em>00 couple-years of follow-up. The upper 95% confidence limit for condomless anal sex was 0.7<em>1</em> per <em>1</em>00 couple-years of follow-up.

CONCLUSIONS

Among serodifferent heterosexual and MSM couples in which the HIV-positive partner was using suppressive ART and who reported condomless sex, during median follow-up of <em>1</em>.3 years per couple, there were no documented cases of within-couple HIV transmission (upper 95% confidence limit, 0.30/<em>1</em>00 couple-years of follow-up). Additional longer-term follow-up is necessary to provide more precise estimates of risk.

The presence and the role of interleukin <em>1</em>0 (IL-<em>1</em>0), a potent cytokine synthesis inhibitory factor and antiinflammatory cytokine, were investigated in rheumatoid arthritis (RA). The expression of both mRNA and protein for IL-<em>1</em>0 could be demonstrated in RA and osteoarthritis (OA) joints. Human IL-<em>1</em>0 mRNA could be demonstrated by polymerase chain reaction amplification of cDNA made by reverse transcription of total RNA extracted directly from synovial tissue in five out of five RA and four out of five OA patients. IL-<em>1</em>0 protein was demonstrated by specific immunoassay and immunohistology. IL-<em>1</em>0 protein was spontaneously produced in all <em>1</em><em>1</em> RA and <em>1</em>7 OA synovial membrane cultures investigated, and this production was sustained for up to 5 d in culture in the absence of any extrinsic stimulation. IL-<em>1</em>0 protein could also be detected by immunohistology in all five RA and four OA synovial membrane biopsies investigated, but not three normal synovial membranes. Immunohistology revealed that the IL-<em>1</em>0 was localized to the synovial membrane lining layer and mononuclear cell aggregates. Immunofluorescence double staining revealed that the sources of IL-<em>1</em>0 were monocytes in the lining layer, and T cells in the mononuclear cell aggregates. We found evidence that the IL-<em>1</em>0 expression was functionally relevant, as neutralization of endogenously produced IL-<em>1</em>0 in the RA synovial membrane cultures resulted in a two- to threefold increase in the protein levels of proinflammatory cytokines tumor necrosis factor alpha (TNF-alpha) and IL-<em>1</em> beta, although IL-6 and IL-8 levels were not affected. The addition of exogenous recombinant IL-<em>1</em>0 to the RA synovial membrane cultures resulted in a two- to threefold decrease in the levels of TNF-alpha and IL-<em>1</em> beta. IL-8 levels were reduced by day 5; however, IL-6 levels were not affected by exogenous IL-<em>1</em>0. Neutralization of the endogenous IL-<em>1</em>0 in two out of seven RA synovial membrane cultures resulted in the expression of detectable levels of interferon gamma (56<em>1</em>-<em>1</em>,050 pg/<em>ml</em>). Taken together, the above findings suggest that IL-<em>1</em>0 is spontaneously produced in RA and OA and is an important immunoregulatory component in the cytokine network of RA, regulating monocyte and in some cases T cell cytokine production.

Angiogenesis is important in the proliferation of inflammatory synovial tissue. Vascular endothelial growth factor (VEGF) is an endothelial cell mitogen that is also angiogenic in vivo. We examined the potential role of VEGF in mediating chemotaxis and proliferation of endothelial cells in rheumatoid arthritis (RA) using samples of synovial tissue and synovial fluid from 55 arthritic patients. Synovial fluid VEGF by ELISA was higher in RA synovial fluids (386 +/- <em>1</em>22 ng/<em>ml</em>) (SE) compared with osteoarthritis (OA) synovial fluids (< 0.8 ng/<em>ml</em>) (p < 0.05) or synovial fluids from patients with other arthritides (6.6 +/- 2 ng/<em>ml</em>). In addition to its known mitogenic properties, we found that human rVEGF was chemotactic for HUVECs at concentrations above 0.02 nM. Incubation of RA synovial fluids with neutralizing anti-VEGF resulted in 23 to 66% (mean 53 +/- 4%) inhibition of HUVEC chemotaxis. Conditioned medium from four of five RA synovial tissue explants was mitogenic for bovine adrenal capillary endothelial cells. Anti-VEGF neutralized from <em>1</em>9 to 42% (mean 28 +/- 4%) of this mitogenic activity. To determine the cellular source of VEGF in synovial tissue, we employed immunohistochemistry. VEGF+ cells were rarely (< <em>1</em>%+) found in normal synovial tissues. In contrast, RA and OA synovial tissues exhibited VEGF+ lining cells (8% and <em>1</em>3%, respectively). A few synovial tissue macrophages were VEGF+ in both RA and OA (5% and 2%, respectively). These results elucidate a newly described function for VEGF as a potent chemotaxin for endothelial cells as well as a role for VEGF in RA-associated endothelial migration and proliferation.

Proinflammatory cytokines and serotonergic homeostasis have both been implicated in the pathophysiology of major psychiatric disorders. We have demonstrated that activation of p38 mitogen-activated protein kinase (MAPK) induces a catalytic activation of the serotonin transporter (SERT) arising from a reduction in the SERT Km for 5-hydroxytryptamine (5-HT). As inflammatory cytokines can activate p38 MAPK, we hypothesized that they might also activate neuronal SERT. Indeed, Interleukin-<em>1</em>beta (IL-<em>1</em>beta) and tumor necrosis factor alpha (TNF-alpha) stimulated serotonin uptake in both the rat embryonic raphe cell line, RN46A, and in mouse midbrain and striatal synaptosomes. In RN46A cells, IL-<em>1</em>beta stimulated 5-HT uptake in a dose- and time-dependent manner, peaking in 20 min at <em>1</em>00 ng/<em>ml</em>. This was abolished by IL-<em>1</em>ra (20 ng/<em>ml</em>), an antagonist of the IL-<em>1</em> receptor, and by SB203580 (5 microM), a p38 MAPK inhibitor. TNF-alpha also dose- and time-dependently stimulated 5-HT uptake that was only partially blocked by SB203580. Western blots showed that IL-<em>1</em>beta and TNF-alpha activated p38 MAPK, in an SB203580-sensitive manner. IL-<em>1</em>beta induced an SB203580-sensitive decrease in 5-HT Km with no significant change in Vmax. In contrast, TNF-alpha stimulation decreased 5-HT Km and increased SERT Vmax. SB203580 selectively blocked the TNF-alpha-induced change in SERT Km. In mouse midbrain and striatal synaptosomes, maximal stimulatory effects on 5-HT uptake occurred at lower concentrations (IL-<em>1</em>beta, <em>1</em>0 ng/<em>ml</em>; TNF-alpha, 20 ng/<em>ml</em>), and over shorter incubation times (<em>1</em>0 min). As with RN46A cells, the effects of IL-<em>1</em>beta and TNF-alpha were completely (IL-<em>1</em>beta) or partially (TNF-alpha) blocked by SB203580. These results provide the first evidence that proinflammatory cytokines can acutely regulate neuronal SERT activity via p38 MAPK-linked pathways.

OBJECTIVE

β-Blocker therapy may control heart rate and attenuate the deleterious effects of β-adrenergic receptor stimulation in septic shock. However, β-Blockers are not traditionally used for this condition and may worsen cardiovascular decompensation related through negative inotropic and hypotensive effects.

OBJECTIVE

To investigate the effect of the short-acting β-blocker esmolol in patients with severe septic shock.

METHODS

Open-label, randomized phase 2 study, conducted in a university hospital intensive care unit (ICU) between November 20<em>1</em>0 and July 20<em>1</em>2, involving patients in septic shock with a heart rate of 95/min or higher requiring high-dose norepinephrine to maintain a mean arterial pressure of 65 mm Hg or higher.

METHODS

We randomly assigned 77 patients to receive a continuous infusion of esmolol titrated to maintain heart rate between 80/min and 94/min for their ICU stay and 77 patients to standard treatment.

METHODS

Our primary outcome was a reduction in heart rate below the predefined threshold of 95/min and to maintain heart rate between 80/min and 94/min by esmolol treatment over a 96-hour period. Secondary outcomes included hemodynamic and organ function measures; norepinephrine dosages at 24, 48, 72, and 96 hours; and adverse events and mortality occurring within 28 days after randomization.

RESULTS

Targeted heart rates were achieved in all patients in the esmolol group compared with those in the control group. The median AUC for heart rate during the first 96 hours was -28/min (IQR, -37 to -2<em>1</em>) for the esmolol group vs -6/min (95% CI, -<em>1</em>4 to 0) for the control group with a mean reduction of <em>1</em>8/min (P < .00<em>1</em>). For stroke volume index, the median AUC for esmolol was 4 <em>mL</em>/m2 (IQR, -<em>1</em> to <em>1</em>0) vs <em>1</em> <em>mL</em>/m2 for the control group (IQR, -3 to 5; P = .02), whereas the left ventricular stroke work index for esmolol was 3 <em>mL</em>/m2 (IQR, 0 to 8) vs <em>1</em> <em>mL</em>/m2 for the control group (IQR, -2 to 5; P = .03). For arterial lactatemia, median AUC for esmolol was -0.<em>1</em> mmol/L (IQR, -0.6 to 0.2) vs 0.<em>1</em> mmol/L for the control group (IQR, -0.3 for 0.6; P = .007); for norepinephrine, -0.<em>1</em><em>1</em> μg/kg/min (IQR, -0.46 to 0.02) for the esmolol group vs -0.0<em>1</em> μg/kg/min (IQR, -0.2 to 0.44) for the control group (P = .003). Fluid requirements were reduced in the esmolol group: median AUC was 3975 <em>mL</em>/24 h (IQR, 3663 to 4200) vs 4425 <em>mL</em>/24 h(IQR, 4038 to 4775) for the control group (P < .00<em>1</em>). We found no clinically relevant differences between groups in other cardiopulmonary variables nor in rescue therapy requirements. Twenty-eight day mortality was 49.4% in the esmolol group vs 80.5% in the control group (adjusted hazard ratio, 0.39; 95% CI, 0.26 to 0.59; P < .00<em>1</em>).

CONCLUSIONS

For patients in septic shock, open-label use of esmolol vs standard care was associated with reductions in heart rates to achieve target levels, without increased adverse events. The observed improvement in mortality and other secondary clinical outcomes warrants further investigation.

BACKGROUND

clinicaltrials.gov Identifier: NCT0<em>1</em>23<em>1</em>698.

TNF-alpha plays a central role in the intestinal inflammation of various inflammatory disorders including Crohn's disease (CD). TNF-alpha-induced increase in intestinal epithelial tight junction (TJ) permeability has been proposed as one of the proinflammatory mechanisms contributing to the intestinal inflammation. The intracellular mechanisms involved in the TNF-alpha-induced increase in intestinal TJ permeability remain unclear. The purpose of this study was to investigate the possibility that the TNF-alpha-induced increase in intestinal epithelial TJ permeability was regulated by myosin light-chain kinase (MLCK) protein expression, using an in vitro intestinal epithelial model system consisting of the filter-grown Caco-2 intestinal epithelial monolayers. TNF-alpha (<em>1</em>0 ng/<em>ml</em>) produced a time-dependent increase in Caco-2 MLCK expression. The TNF-alpha increase in MLCK protein expression paralleled the increase in Caco-2 TJ permeability, and the inhibition of the TNF-alpha-induced MLCK expression (by cycloheximide) prevented the increase in Caco-2 TJ permeability, suggesting that MLCK expression may be required for the increase in Caco-2 TJ permeability. The TNF-alpha increase in MLCK protein expression was preceded by an increase in MLCK mRNA expression but not an alteration in MLCK protein degradation. Actinomycin-D prevented the TNF-alpha increase in MLCK mRNA expression and the subsequent increase in MLCK protein expression and Caco-2 TJ permeability, suggesting that the increase in MLCK mRNA transcription led to the increase in MLCK expression. The TNF-alpha increase in MLCK protein expression was also associated with an increase in Caco-2 MLCK activity. The cycloheximide inhibition of MLCK protein expression prevented the TNF-alpha increase in MLCK activity and Caco-2 TJ permeability. Moreover, inhibitors of MLCK, Mg(2+)-myosin ATPase, and metabolic energy prevented the TNF-alpha increase in Caco-2 TJ permeability, suggesting that the increase in MLCK activity was required for the TNF-alpha-induced opening of the Caco-2 TJ barrier. In conclusion, our results indicate for the first time that <em>1</em>) the TNF-alpha increase in Caco-2 TJ permeability was mediated by an increase in MLCK protein expression, 2) the increase in MLCK protein expression was regulated by an increase in MLCK mRNA transcription, and 3) the increase in Caco-2 TJ permeability required MLCK protein expression-dependent increase in MLCK activity.

We have investigated an improved method for generating sizable numbers of mature dendritic cells from nonproliferating progenitors in human blood. The procedure uses <em>1</em>% human plasma in the place of <em>1</em>0% fetal calf serum and involves two steps. The first step or 'priming' phase is a 6-7 day culture of T cell depleted mononuclear cells in medium supplemented with GM-CSF and IL-4. The second step or 'differentiation' phase requires the exposure to macrophage conditioned medium. This medium cannot be replaced by several known cytokines such as TNF-alpha, IL-<em>1</em>, IL-6, IL-<em>1</em>2 and IL-<em>1</em>5, and cannot be inhibited with neutralizing antibodies to IL-<em>1</em>, TNF-alpha, IL-6 or IL-<em>1</em>2 alone, or in combination. Using this two-step approach, we obtain substantial yields. About <em>1</em>-3 x <em>1</em>0(6) mature dendritic cells are generated from 40 <em>ml</em> of blood vs. < 0.<em>1</em> x <em>1</em>0(6) from noncytokine treated blood. The dendritic cells derive from progenitors found primarily in a radioresistant population of CD<em>1</em>4+ and adherent blood mononuclear cells and have all the features of mature cells. They include a stellate cell shape, nonadherence to plastic, and very strong T cell stimulatory activity. Strong APC function was evident for both the proliferation of allogeneic T cells in the MLR, and the generation by syngeneic T cells of class I restricted, CTL responses to influenza virus. A panel of dendritic cell restricted markers is also expressed, including CD83, p55, and perinuclear CD68. All of these dendritic cell properties are retained for at least 3 days when the cytokines are removed, suggesting that these populations are stable and terminally differentiated. We suggest that these cells will be effective in vivo as adjuvants for active immunotherapy.

BACKGROUND

The gut is a major reservoir for human immunodeficiency virus (HIV) in patients receiving antiretroviral therapy (ART). We hypothesized that distinct immune environments within the gut may support varying levels of HIV.

METHODS

In 8 HIV-<em>1</em>-positive adults who were receiving ART and had CD4(+) T cell counts of >200 cells/<em>μL</em> and plasma viral loads of <40 copies/<em>mL</em>, levels of HIV and T cell activation were measured in blood samples and endoscopic biopsy specimens from the duodenum, ileum, ascending colon, and rectum.

RESULTS

HIV DNA and RNA levels per CD4(+) T cell were higher in all 4 gut sites compared with those in the blood. HIV DNA levels increased from the duodenum to the rectum, whereas the median HIV RNA level peaked in the ileum. HIV DNA levels correlated positively with T cell activation markers in peripheral blood mononuclear cells (PBMCs) but negatively with T cell activation markers in the gut. Multiply spliced RNA was infrequently detected in gut, and ratios of unspliced RNA to DNA were lower in the colon and rectum than in PBMCs, which reflects paradoxically low HIV transcription, given the higher level of T cell activation in the gut.

CONCLUSIONS

HIV DNA and RNA are both concentrated in the gut, but the inverse relationship between HIV DNA levels and T cell activation in the gut and the paradoxically low levels of HIV expression in the large bowel suggest that different processes drive HIV persistence in the blood and gut.

BACKGROUND

ClinicalTrials.gov identifier: NCT00884793 (PLUS<em>1</em>).

OBJECTIVE

To determine utility of multiparametric imaging performed at 3 T for detection of prostate cancer by using T2-weighted magnetic resonance (MR) imaging, MR spectroscopy, and dynamic contrast material-enhanced MR imaging, with whole-mount pathologic findings as reference standard.

METHODS

This prospectively designed, HIPAA-compliant, single-institution study was approved by the local institutional review board. Seventy consecutive patients (mean age, 60.4 years; mean prostate-specific antigen level, 5.47 ng/<em>mL</em> [5.47 microg/L]; range, <em>1</em>-<em>1</em>9.9 ng/<em>mL</em> [<em>1</em>-<em>1</em>9.9 microg/L]) were included; informed consent was obtained from each patient. All patients had biopsy-proved prostate cancer, with a median Gleason score of 7 (range, 6-9). Images were obtained by using a combination of six-channel cardiac and endorectal coils. MR imaging and pathologic findings were evaluated independently and blinded and then correlated with histopathologic findings by using side-by-side comparison. Analyses were conducted with a raw stringent approach and an alternative neighboring method, which accounted for surgical deformation, shrinkage, and nonuniform slicing factors in pathologic specimens. Generalized estimating equations (GEEs) were used to estimate the predictive value of region-specific, pathologically determined cancer for all three modalities. This approach accounts for the correlation among multiple regions in the same individual.

RESULTS

For T2-weighted MR imaging, sensitivity and specificity values obtained with stringent approach were 0.42 (95% confidence interval [CI]: 0.36, 0.47) and 0.83 (95% CI: 0.8<em>1</em>, 0.86), and for the alternative neighboring approach, sensitivity and specificity values were 0.73 (95% CI: 0.67, 0.78) and 0.89 (95% CI: 0.85, 0.93), respectively. The combined diagnostic accuracy of T2-weighted MR imaging, dynamic contrast-enhanced MR imaging, and MR spectroscopy for peripheral zone tumors was examined by calculating their predictive value with different combinations of techniques; T2-weighted MR imaging, dynamic contrast-enhanced MR imaging, and MR spectroscopy provided significant independent and additive predictive value when GEEs were used (P < .00<em>1</em>, P = .02, P = .002, respectively).

CONCLUSIONS

Multiparametric MR imaging (T2-weighted MR imaging, MR spectroscopy, dynamic contrast-enhanced MR imaging) of the prostate at 3 T enables tumor detection, with reasonable sensitivity and specificity values.

Inflammation plays an important role in the development of atherosclerosis, but the specific stimuli governing cytokine release in atherogenesis are unknown. We examined the hypothesis that hypertension may increase the risk of atherosclerosis via proinflammatory effects. In a cross-sectional study involving 508 apparently healthy men, we studied the association between blood pressure and baseline plasma concentrations of 2 inflammatory markers, intercellular adhesion molecule-<em>1</em> (sICAM-<em>1</em>) and interleukin-6 (IL-6). Increase in systolic blood pressure (SBP) (P=0.003), pulse pressure (PP) (P=0.0<em>1</em>9), and mean arterial pressure (P=0.0<em>1</em>4) was significantly associated with levels of sICAM-<em>1</em>. All of these measures of blood pressure, as well as diastolic blood pressure (DBP), were significantly associated with levels of IL-6 (all, P</=0.00<em>1</em>). In multiple linear regression models controlled for age and other cardiac risk factors, SBP (7.6 ng/<em>mL</em> per <em>1</em>0 mm Hg, P=0.0<em>1</em>6) and PP (8.<em>1</em>3 ng/<em>mL</em> per <em>1</em>0 mm Hg, P=0.038) were significantly associated with sICAM-<em>1</em> levels, whereas SBP (0.<em>1</em><em>1</em> pg/<em>mL</em> per <em>1</em>0 mm Hg, P<0.00<em>1</em>), DBP (0.<em>1</em><em>1</em> pg/<em>mL</em> per <em>1</em>0 mm Hg, P=0.008), PP (0.<em>1</em>0 pg/<em>mL</em> per <em>1</em>0 mm Hg, P=0.009), and mean arterial pressure (0.<em>1</em>5 pg/<em>mL</em> per <em>1</em>0 mm Hg, P<0.00<em>1</em>) had similar strong relationships with log-transformed IL-6 levels. Therefore, in apparently healthy men, we observed significant graded relationships between blood pressure and levels of sICAM-<em>1</em> as well as IL-6. These data suggest that increased blood pressure may be a stimulus for inflammation and that this is a possible mechanism underlying the well-established role of hypertension as a risk factor for atherosclerotic disease.

BACKGROUND

Substantial morbidity occurs during the first year of antiretroviral therapy (ART) in persons with advanced human immunodeficiency virus (HIV) disease despite HIV suppression. Biomarkers may identify high-risk groups.

METHODS

Pre-ART and <em>1</em>-month samples from an initial ART trial were evaluated for biomarkers associated with AIDS events or death within <em>1</em>-<em>1</em>2 months. Case patients (n = 63) and control patients (n = <em>1</em>26) were <em>1</em>:2 matched on baseline CD4 cell count, hepatitis status, and randomization date. All had ≥ <em>1</em> log(<em>1</em>0) HIV RNA level decrease at <em>1</em> month.

RESULTS

Case patients had more frequent prior AIDS events, compared with control patients (P = .004), but similar HIV RNA levels at baseline. Pre-ART and <em>1</em>-month C-reactive protein (CRP), D-dimer, and interleukin 6 (IL-6) levels and pre-ART hyaluronic acid (HA) levels were associated with new AIDS events or death (P ≤ .0<em>1</em>). Patients who experienced immune reconstitution inflammatory syndrome (IRIS) events had higher pre-ART tumor necrosis factor α (TNF-α) and HIV RNA levels and significant <em>1</em>-month increases in CRP, D-dimer, IL-6, interleukin 8, CXCL<em>1</em>0, TNF-α, and interferon-γ levels, compared with patients who experienced non-IRIS events (P ≤ .03). Individuals with baseline CRP and HA levels above the cohort median (>2.<em>1</em> mg/L and >50.0 ng/mL, respectively) had increased risk of AIDS or death (OR, 4.6 [95% CI, 2.0-<em>1</em>0.3]; P < .00<em>1</em>) and IRIS (OR, 8.7 [95% CI, 2.2-34.8] P = .002).

CONCLUSIONS

Biomarkers of Inflammation (CRP, IL-6), coagulation (D-dimer), and tissue fibrosis (HA) measured pre-ART and at <em>1</em> month are associated with higher risk of AIDS events, IRIS, or death, warranting additional study as risk stratification strategies.

Human recombinant tumor necrosis factor (TNF) induced migration across polycarbonate and nitrocellulose filters of human peripheral blood monocytes and polymorphonuclear leukocytes, TNF was active in inducing migration at concentrations less than <em>1</em> U/<em>ml</em>, and maximal responses (observed at greater than <em>1</em>00 U/<em>ml</em>) were comparable to those elicited by standard reference chemoattractants (FMLP, <em>1</em>0 nM; activated human serum, 5%). Checkerboard analysis performed by seeding different concentrations of TNF above and below the filter revealed that maximal induction of migration required a positive concentration gradient between the lower and upper compartments and that TNF elicited an actual chemotactic response in phagocytes. An anti-TNF rabbit antiserum and anti-TNF mouse monoclonal antibody abolished the chemotactic activity of TNF. Recombinant lymphotoxin was also chemotactic for phagocytes, and its activity was blocked by an anti-lymphotoxin antiserum. Human umbilical vein endothelial cells and blood large granular lymphocytes did not respond chemotactically to TNF under conditions in which appropriate reference chemoattractants were active. The chemotactic activity of TNF may serve to recruit phagocytic cells from the blood compartment to amplify resistance against noxious agents.