Bridging barriers to clinic-based HIV testing with new technology: translating self-implemented testing for African American youth.

Numerous barriers to clinic-based HIV testing exist (e.g., stigmatization) for African American youth. These barriers may be addressed by new technology, specifically HIV self-implemented testing (SIT). We conducted a series of formative phase 3 translation studies (49 face-to-face interviews, 9 focus groups, 1 advisory panel review) among low-income African American youth (15-19 years) and providers of adolescent services in two US cities to identify potential translation difficulties of the OraQuick SIT. Based on content analysis, we found that providers and African American youth viewed SITs positively compared to clinic-based testing. Data suggest that SITs may reduce social stigma and privacy concerns and increase convenience and normalization of HIV testing. Challenges with SIT implementation include difficulties accessing confirmatory testing, coping with adverse outcomes, and instructional materials that may be inappropriate for low socioeconomic status (SES) persons. Study results underscore the need for translation studies to identify specific comprehension and implementation problems African American youth may have with oral SITs.

Increased susceptibility of aging kidney to ischemic injury: identification of candidate genes changed during aging, but corrected by caloric restriction.

Aging is associated with an increased incidence and severity of acute renal failure. However, the molecular mechanism underlying the increased susceptibility to injury remains undefined. These experiments were designed to investigate the influence of age on the response of the kidney to ischemic injury and to identify candidate genes that may mediate this response. Renal slices prepared from young (5 mo), aged ad libitum (aged-AL; 24 mo), and aged caloric-restricted (aged-CR; 24 mo) male Fischer 344 rats were subjected to ischemic stress (100% N(2)) for 0-60 min. As assessed by biochemical and histological evaluation, slices from aged-AL rats were more susceptible to injury than young counterparts. Importantly, caloric restriction attenuated the increased susceptibility to injury. In an attempt to identify the molecular pathway(s) underlying this response, microarray analysis was performed on tissue harvested from the same animals used for the viability experiments. RNA was isolated and the corresponding cDNA was hybridized to CodeLink Rat Whole Genome Bioarray slides. Subsequent gene expression analysis was performed using GeneSpring software. Using two-sample t-tests and a twofold cut-off, the expression of 92 genes was changed during aging and attenuated by caloric restriction, including claudin-7, kidney injury molecule-1 (Kim-1), and matrix metalloproteinase-7 (MMP-7). Claudin-7 gene expression peaked at 18 mo; however, increased protein expression in certain tubular epithelial cells was seen at 24 mo. Kim-1 gene expression was not elevated at 8 or 12 mo but was at 18 and 24 mo. However, changes in Kim-1 protein expression were only seen at 24 mo and corresponded to increased urinary levels. Importantly, these changes were attenuated by caloric restriction. MMP-7 gene expression was decreased at 8 mo, but an age-dependent increase was seen at 24 mo. Increased MMP-7 protein expression in tubular epithelial cells at 24 mo was correlated with the gene expression pattern. In summary, we identified genes changed by aging and changes attenuated by caloric restriction. This will facilitate investigation into the molecular mechanism mediating the age-related increase in susceptibility to injury.

Precision-cut tissue chips as an in vitro toxicology system.

Precision-cut tissue slices mimic specific organ toxicity because normal cellular heterogeneity and organ architecture are retained. To optimize the use of the smaller tissues of the mouse and to establish easy assays for tissue viability, a tissue chip based system was used to generate large numbers of samples from a single organ. Iodoacetamide (IAM) was used as a model toxicant and assays for intracellular potassium (normalized to DNA content) were used to establish viability and toxicant susceptibility. Thereafter, assays that were more rapid and specific were pursued. Lysates from tissues incubated in 6-carboxyfluorescein fluoresced proportionately to concentrations of IAM, indicating disruption of cellular membranes. Similarly, FURA-2, a probe applied to lysates to measure calcium levels, fluoresced proportionately to IAM dosage. Monobromobimane, a fluorescent sulfhydryl probe, displayed a decrease in fluorescent intensity at higher IAM challenge-a finding confirmed with an absorbance assay with Ellman's reagent. Importantly, the number of samples per organ/mouse was increased at least threefold and a significant time reduction per analysis was realized.

Role of matrix metalloproteinases in renal pathophysiologies.

Matrix metalloproteinases (MMPs) are a large family of proteinases that remodel extracellular matrix (ECM) components and cleave a number of cell surface proteins. MMP activity is regulated via a number of mechanisms, including inhibition by tissue inhibitors of metalloproteinases (TIMPs). Originally thought to cleave only ECM proteins, MMP substrates are now known to include signaling molecules (growth factor receptors) and cell adhesion molecules. Recent data suggest a role for MMPs in a number of renal pathophysiologies, both acute and chronic. This review will focus on the expression and localization of MMPs and TIMPs in the kidney, as well as summarizing the current information linking these proteins to acute kidney injury, glomerulosclerosis/tubulointerstitial fibrosis, chronic allograft nephropathy, diabetic nephropathy, polycystic kidney disease, and renal cell carcinoma.

Low-level arsenite causes accumulation of ubiquitinated proteins in rabbit renal cortical slices and HEK293 cells.

Arsenic is a known human carcinogen that affects a variety of processes within the cell. In this study, the effects of environmentally relevant As(III) exposures on the ubiquitin (Ub)-proteasome pathway have been investigated. Low-level As(III) exposure (0.5 - 10 microM) causes an accumulation of high-molecular-weight ubiquitin protein conjugates in both precision-cut rabbit renal-cortical slices and human embryonic kidney (HEK) 293 cells. The As(III) doses that induced these molecular changes were subcytotoxic in both model systems. Doses of 10 microM As(III) decreased cellular activity of the 20S proteasome by 40 and 15% in slices and HEK293 cells, respectively. As(III) did not cause any notable difference in Ub-conjugating activity of rabbit renal slices or HEK293 cells. Since ubiquitination plays such a vital role in maintaining cellular homeostasis, this noticeable perturbation of cellular ubiquitination is likely to have a multitude of signaling effects within the cells and may contribute to the pathogenesis of low-level arsenic.

Precision-cut tissue slices from transgenic mice as an in vitro toxicology system.

In these experiments precision-cut tissue slices from two existing transgenic mouse strains, with transgenes that couple promoting or binding elements to a reporter protein, were used for determination of reporter induction. This approach combines the power of transgenic animals with the practicality of in vitro systems to investigate the biological impact of xenobiotics. Additionally, the normal cellular architecture and heterogeneity is retained in precision-cut tissue slices. Two transgenic mouse strains, one of which couples the promoting region of CYP 1A1 to beta-galactosidase, and another which couples two forward and two backward 12-O-tetradecanoyl phorbol-13-acetate (TPA) repeat elements (TRE) to luciferase (termed AP-1/luciferase), were used to determine the feasibility of this approach. Precision-cut kidney and liver slices from both transgenic strains remain viable as determined by slice K(+) ion content and LDH enzyme release. Liver slices harvested from the CYP 1A1/beta-galactosidase transgenic mice exhibit a 14-fold increase in beta-galactosidase activity when incubated with beta-napthoflavone for 24 h. Kidney and liver slices obtained from the AP-1/luciferase transgenic mice demonstrate induction of luciferase (up to 2.5-fold) when incubated with phorbol myristate acetate (PMA or TPA) up to 4 h. These data indicate that precision-cut tissue slices from transgenic mice offer a novel in vitro method for toxicity evaluation while maintaining normal cell heterogeneity.

The role of acetylation in rDNA transcription.

Treatment of NIH 3T3 cells with trichostatin A (TSA), an inhibitor of histone deacetylase (HDAC), resulted in a dose-dependent increase in transcription from a rDNA reporter and from endogenous rRNA genes. Chromatin immunoprecipitation using anti-acetyl-histone H4 antibodies demonstrated a direct effect of TSA on the acetylation state of the ribosomal chromatin. TSA did not reverse inhibition of transcription from the rDNA reporter by retinoblastoma (Rb) protein, suggesting that the main mechanism by which Rb blocks rDNA transcription may not involve recruitment of deacetylases to rDNA chromatin. Overexpression of histone transacetylases p300, CBP and PCAF stimulated transcription in transfected NIH 3T3 cells. Recombinant p300, but not PCAF, stimulated rDNA transcription in vitro in the absence of nucleosomes, suggesting that the stimulation of rDNA transcription by TSA might have a chromatin-independent component. We found that the rDNA transcription factor UBF was acetylated in vivo. Finally, we also demonstrated the nucleolar localization of CBP. Our results suggest that the organization of ribosomal chromatin of higher eukaryotes is not static and that acetylation may be involved in affecting these dynamic changes directly through histone acetylation and/or through acetylation of UBF or one of the other components of rDNA transcription.

Toxicity of a sevoflurane degradation product incubated with rat liver and renal cortical slices.

Compound A (2-fluoromethoxy-1,1,3,3,3-pentafluoro-1-propene) is a degradation product of the anesthetic sevoflurane which is created in closed-circuit anesthetic machines. Past in vivo and in vitro studies have implied that Compound A is nephrotoxic via bioactivation through the cysteine conjugate beta-lyase pathway. Although glutathione (GSH) conjugates of Compound A have been reported, it is not clear if they are formed enzymatically or via direct reaction with GSH. To determine if these metabolites are produced and toxic, a tissue slice system that first exposes male Fischer 344 rat liver slices to volatilized Compound A followed by exposure of rat kidney slices to the liver incubate was employed. Liver slices exposed to volatilized Compound A (6-12 microM medium conc.; approximately 23 ppm) exhibited a loss of K+ by 6 h, which was not seen in kidney slices exposed to Compound A. Aminobenzotriazole, a cytochrome P 450 suicide inhibitor, initially inhibits the cytotoxicity of Compound A to liver slices (at these times and concentrations). The sequential liver/kidney slice experiments using Compound A have not demonstrated nephrotoxic results. GSH conjugates were synthesized and was found to be nephrotoxic at concentrations above 91 microM (18 h), with higher concentrations showing toxicity at earlier times. Additionally, non-enzymatic reactions of Compound A with GSH or sulfhydryl-containing medium produces nephrotoxic products. These studies show that Compound A is directly toxic to the liver, possibly via P 450 activation, and Compound A can react with sulfhydryls directly to produce a nephrotoxic.

Differential HIV risk in bathhouses and public cruising areas.

OBJECTIVES: This report investigates differences in risk behaviors among men who have sex with men (MSM) who went to gay bathhouses, public cruising areas, or both. METHODS: We used a probability sample of MSM residing in 4 US cities (n = 2,881). RESULTS: Men who used party drugs and had unprotected anal intercourse with nonprimary partners were more likely to go to sex venues than men who did not. Among attendees, MSM who went to public cruising areas only were least likely, and those who went to both public cruising areas and bathhouses were most likely to report risky sex in public settings. CONCLUSIONS: Distinguishing between sex venues previously treated as a single construct revealed a significant association between pattern of venue use and sexual risk. Targeting HIV prevention in the bathhouses would reach the segment of men at greatest risk for HIV transmission.

The continuing HIV epidemic among men who have sex with men.

OBJECTIVES: This study characterized the AIDS epidemic among urban men who have sex with men (MSM). METHODS: A probability sample of MSM was obtained in 1997 (n = 2881; 18 years and older) from New York, Los Angeles, Chicago, and San Francisco, and HIV status was determined through self-report and biological measures. RESULTS: HIV prevalence was 17% (95% confidence interval = 15%, 19%) overall, with extremely high levels in African Americans (29%), MSM who used injection drugs (40%), "ultraheavy" noninjection drug users (32%), and less educated men (< high school, 37%). City-level HIV differences were non-significant once these other factors were controlled for. In comparing the present findings with historical data based on public records and modeling, HIV prevalence appears to have declined as a result of high mortality (69%) and stable, but high, incidence rates (1%-2%). CONCLUSIONS: Although the findings suggest that HIV prevalence has declined significantly from the mid-1980s, current levels among urban MSM in the United States approximate those of sub-Saharan countries (e.g., 14%-25%) and are extremely high in many population subsegments. Despite years of progress, the AIDS epidemic continues unabated among subsegments of the MSM community.

Health-related characteristics of men who have sex with men: a comparison of those living in "gay ghettos" with those living elsewhere.

OBJECTIVES: This study investigated the limitations of probability samples of men who have sex with men (MSM), limited to single cities and to the areas of highest concentrations of MSM ("gay ghettos"). METHODS: A probability sample of 2881 MSM in 4 American cities completed interviews by telephone. RESULTS: MSM who resided in ghettos differed from other MSM, although in different ways in each city. Non-ghetto-dwelling MSM were less involved in the gay and lesbian community. They were also less likely to have only male sexual partners, to identify as gay, and to have been tested for HIV. CONCLUSIONS: These differences between MSM who live in gay ghettos and those who live elsewhere have clear implications for HIV prevention efforts and health care planning.

National trends in condom use among at-risk heterosexuals in the united states.

Based on national level surveys, we examined data relevant to the United States' overall effort to prevent the spread of HIV among heterosexual adults. We examined changes in condom use among at-risk heterosexuals over the past decade. The observed increases over time in condom use across all heterosexual at-risk population segments are consistent with the observed (declines) trends in HIV and syphilis in the 1990s. These results and findings from prior studies suggest that U.S. efforts to facilitate condom use and contain HIV and related sexually transmitted disease (STD)-cofactors among adult at risk heterosexuals was succeeding over most of the 1990s. The absence of national level behavioral trend data after 1996, and the ambiguities of HIV spread suggest some caution in projecting trends into this century. National and local efforts need to be directed at sustaining behavioral change and conducting more rigorous studies on population trends in HIV/STD-related behaviors/pathogens.

Use of antiretroviral therapies among HIV-infected men who have sex with men: a household-based sample of 4 major American cities.

OBJECTIVES: This study sought to determine the prevalence and determinants of use of recommended antiretroviral regimens among urban seropositive men who have sex with men (MSM). METHODS: A probability telephone sample of MSM was taken within regions of Chicago, Los Angeles, New York, and San Francisco. Analysis focused on use of antiretroviral therapies. RESULTS: Although the majority of seropositive MSM with CD4 counts below 500 per microliter were using recommended antiretroviral regimens, 26% of seropositive MSM were not receiving such care. Men who were younger, who reported a sexual orientation other than homosexual, who had a more recent interview date, who were at middle levels of affiliation with the gay community, and who reported higher levels of perceived exclusivity on the part of the gay community were less likely to be using recommended antiretroviral regimens. CONCLUSIONS: Although current efforts to make antiretroviral therapies available to HIV-seropositive MSM are reasonably effective, additional efforts are needed for MSM characterized by relative youth and lower social support.

Understanding childhood sexual abuse as a predictor of sexual risk-taking among men who have sex with men: The Urban Men's Health Study.

OBJECTIVE: The prevalence and characteristics of childhood sexual abuse (CSA) among men who have sex with men (MSM), and links with sexual risk are explored. A model linking CSA and sexual risk among MSM is proposed. METHOD: A telephone probability sample of urban MSM (n = 2881) was recruited and interviewed between November 1996 and February 1998. The interview covered numerous health issues, including history of sexual victimization. RESULTS: One-fifth reported CSA, primarily by non-family perpetrators. Initial CSA experiences are characterized by high levels of force (43% involved physical force/weapons), and penetrative sex (78%; 46% reported attempted or actual anal intercourse). Such men are more likely than nevercoerced men to engage in high risk sex (unprotected anal intercourse with a non-primary partner or with a serodiscordant male). In multivariate analyses, the effect of childhood sexual coercion on sexual risk is mediated by substance use, patterns of sexual contacts, and partner violence, but not by adult sexual revictimization or by depression. CONCLUSIONS: Findings are interpreted within the context of social learning theory and prior research on sexual risk-taking. The high risk for CSA among MSM, which can predispose such men to patterns of HIV sexual risk, warrants new approaches in HIV prevention.

Alcohol use, drug use and alcohol-related problems among men who have sex with men: the Urban Men's Health Study.

AIMS: To measure the prevalence and independent associations of heavy and problematic use of alcohol and recreational drugs among a household-based sample of urban MSM (men who have sex with men). DESIGN: Cross-sectional survey. PARTICIPANTS: Men who identified as being gay or bisexual or who reported sex with another man in the prior 5 years were included in this analysis (n = 2172). SETTING: A probability telephone sample of MSM was taken within Zip Codes of four large American cities (Chicago, Los Angeles, New York and San Francisco) estimated to have total concentrations of at least 4% of all households with one resident MSM. MEASUREMENTS: Standard measures of alcohol use, problems associated with alcohol use, and recreational drug use were administered by trained telephone interviewers. FINDINGS: Both recreational drug (52%) and alcohol use (85%) were highly prevalent among urban MSM, while current levels of multiple drug use (18%), three or more alcohol-related problems (12%), frequent drug use (19%) and heavy-frequent alcohol use (8%) were not uncommon. The associations of heavy and/or problematic substance use are complex, with independent multivariate associations found at the levels of demographics, adverse early life circumstances, current mental health status, social and sexual practices and connection to gay male culture. CONCLUSIONS: The complex pattern of associations with heavy and/or problematic substance use among urban MSM suggests that heavy and/or problematic substance use is grounded in multiple levels: the individual, the interpersonal and the socio-cultural.

Involvement of Rb family proteins, focal adhesion proteins and protein synthesis in senescent morphogenesis induced by hydrogen peroxide.

Early passage human diploid fibroblasts develop senescent morphology prematurely within a week after a 2-hour pulse treatment with low or mild dose H(2)O(2). We test here the role of cell cycle checkpoints, cytoskeletal proteins and de novo protein synthesis in senescent morphogenesis following H(2)O(2) treatment. H(2)O(2) treatment causes transient elevation of p53 protein and prolonged inhibition of Rb hyperphosphorylation. Expression of human papillomaviral E6 gene prevented elevation of p53 but did not affect senescent morphogenesis. Expression of human papillomaviral E7 gene reduced the level of Rb protein and prevented induction of senescent morphology by H(2)O(2). The mutants of the E7 gene, in which the Rb family protein binding site was destroyed, could not reduce Rb protein or prevent H(2)O(2) from inducing senescent morphology. Senescent-like cells showed enhanced actin stress fibers. In untreated cells, vinculin and paxillin preferentially distributed along the edge of the cells. In contrast, vinculin and paxillin distributed randomly and sporadically throughout senescent-like cells. E7 expression prevented enhancement of actin filament formation and redistribution of vinculin or paxillin. Neither wild-type nor E7 cells showed changes in the protein level of actin, vinculin or paxillin measured by western blot after H(2)O(2) treatment. Finally, depletion of methionine in the culture medium after H(2)O(2) treatment prevented senescent morphogenesis without affecting dephosphorylation of Rb protein. Our results suggest that senescent morphology likely develops by a program involving activated Rb family proteins, enhancement of actin stress fibers, redistribution of focal adhesion proteins and de novo protein synthesis.

Obtaining HIV test results with a home collection test kit in a community telephone sample.

OBJECTIVES: To test the feasibility of obtaining HIV test results by home collection kit from a probability telephone sample of men who have sex with men (MSM). METHODS: A quota sample of 615 MSM previously interviewed by the Urban Men's Health Study phone survey in Chicago, Los Angeles, New York City, and San Francisco were re-contacted and offered an HIV test using an oral specimen (Orasure) home collection kit. RESULTS: Eighty percent consented to be mailed a kit, and 84% returned a specimen, for a 67% participation rate. All self-reported HIV-positive persons tested positive (77 of 77); 4 of 266 (1.5%) with a prior negative test and 2 of 69 (2.9%) with no prior positive HIV test result. Participation was associated with self-reported prior HIV test status-HIV-positive (83%), HIV-negative (68%), or no prior HIV test result (54%)-and marginally associated with New York City residence after adjustment for HIV status (odds ratio = 0.7; 95% confidence interval, 0.4-1.1; p =.08). CONCLUSIONS: These results suggest that urban MSM identified and interviewed by telephone will participate in home collection HIV testing. This methodology could be used to produce population-based estimates of HIV seroprevalence and seroincidence in MSM and could probably be extended to other populations and other viral infections.

The impact of T-ACASI interviewing on reported drug use among men who have sex with men.

Measurements of drug use and other illicit or stigmatized behaviors are subject to nontrivial underreporting biases. During in-person surveys, respondents are more likely to report such behaviors when interviewed using techniques that maximize interviewee privacy, e.g., use of paper SAQs and audio-CASI rather than questioning by human interviewers. Until recently, respondents in telephone surveys could not be offered similar privacy. A new technology, telephone audio computer-assisted self-interviewing (T-ACASI) overcomes this limitation of telephone surveys by allowing respondents to respond to a computer. A randomized experimental test of T-ACASI was embedded in the Urban Men's Health Study (UMHS). UMHS surveyed a probability sample of 2,881 men from four United States cities and who reported having sex with men. Respondents interviewed using T-ACASI reported a higher prevalence of drug use and drug-related behaviors than respondents interviewed by human interviewers. However, survey respondents were more likely to break off an interview when the interview was conducted by a T-ACASI computer rather than by a human interviewer.