Motivation and context of concurrent stimulant and opioid use among persons who use drugs in the rural United States: a multi-site qualitative inquiry.

BACKGROUND: In recent years, stimulant use has increased among persons who use opioids in the rural U.S., leading to high rates of overdose and death. We sought to understand motivations and contexts for stimulant use among persons who use opioids in a large, geographically diverse sample of persons who use drugs (PWUD) in the rural settings. METHODS: We conducted semi-structured individual interviews with PWUD at 8 U.S. sites spanning 10 states and 65 counties. Content areas included general substance use, injection drug use, changes in drug use, and harm reduction practices. We used an iterative open-coding process to comprehensively itemize and categorize content shared by participants related to concurrent use. RESULTS: We interviewed 349 PWUD (64% male, mean age 36). Of those discussing current use of stimulants in the context of opioid use (n = 137, 39%), the stimulant most used was methamphetamine (78%) followed by cocaine/crack (26%). Motivations for co-use included: 1) change in drug markets and cost considerations; 2) recreational goals, e.g., seeking stronger effects after heightened opioid tolerance; 3) practical goals, such as a desire to balance or alleviate the effects of the other drug, including the use of stimulants to avoid/reverse opioid overdose, and/or control symptoms of opioid withdrawal; and 4) functional goals, such as being simultaneously energized and pain-free in order to remain productive for employment. CONCLUSION: In a rural U.S. cohort of PWUD, use of both stimulants and opioids was highly prevalent. Reasons for dual use found in the rural context compared to urban studies included changes in drug availability, functional/productivity goals, and the use of methamphetamine to offset opioid overdose. Education efforts and harm reduction services and treatment, such as access to naloxone, fentanyl test strips, and accessible drug treatment for combined opioid and stimulant use, are urgently needed in the rural U.S. to reduce overdose and other adverse outcomes.

Clinical impact of treatment timing for chronic hepatitis C infection: a decision model.

Recent advances in the treatment of hepatitis C virus (HCV) infection have led to the availability of both highly efficacious interferon-containing and interferon-sparing regimens. However, the use of such therapies faces restrictions due to high costs. For patients who are medically eligible to receive interferon, the choice between the two will likely be impacted by preferences surrounding interferon, severity of disease, coverage policies and out-of-pocket costs. We developed a decision model to quantify the trade-offs between immediate, interferon-containing therapy and delayed, interferon-free therapy for patients with chronic, genotype 1 HCV infection. We projected the quality-adjusted life expectancy stratified by the presence or absence of cirrhosis for four strategies: (i) no treatment; (ii) immediate, one-time treatment with an interferon-containing regimen; (iii) immediate treatment as above with the opportunity for retreatment in patients who fail to achieve sustained virologic response with interferon-free therapy in 1 year; and (iv) delayed therapy with interferon-free therapy in 1 year. When compared to one-time immediate treatment with the interferon-containing regimen, delayed treatment with the interferon-free regimen in 1 year resulted in longer life expectancy, with a 0.2 quality-adjusted life year (QALY) increase in noncirrhotic patients, and a 1.1 QALY increase in patients with cirrhosis. This superiority in health benefits was lost when wait time for interferon-free therapy was greater than 3-3.2 years. In this modelling analysis, interferon-free therapy resulted in superior health benefits compared to immediate therapy with interferon until wait time exceeded 3-3.2 years. Such data can inform decision-making regarding treatment initiation for HCV as healthcare financing evolves.

Reovirus as a successful ex vivo purging modality for multiple myeloma.

Autologous stem cell rescue (ASCT) following high-dose myeloablative chemotherapy is considered to be a therapeutic option for many multiple myeloma (MM) patients; however relapse post ASCT presents a major challenge. The oncolytic potential of reovirus has been previously demonstrated and is currently undergoing phase I monotherapy clinical trials for MM and phase II/III clinical trials for solid tumors. Here we tested the hypothesis that reovirus can successfully purge MM in a murine model that partially recapitulates human MM. RPMI 8226, MM1S, H929 and U266 human myeloma cell lines were exposed to reovirus and oncolysis was assessed. Apheresis product admixed with MM cells was purged with live reovirus (LV) or dead virus (DV) and purging efficacy was monitored via flow cytometry, reverse transcribed-PCR (RT-PCR) and disease relapse in non obese diabetic/severe combined immune deficient (NOD/SCID) mice. Significant LV purging was seen with MM1S, H929 and U266 and the complete ex vivo purging achieved with RPMI 8226 was confirmed by flow cytometry, RT-PCR and absence of disease relapse in vivo. Mice that received LV-purged autografts exhibited 100% survival in comparison to mice that received DV-purged controls. Reovirus's unique ability to kill MM while sparing hematopoietic stem cells places it as an attractive purging agent for MM during ASCT.

Cofilin is a marker of myofibroblast differentiation in cells from porcine aortic cardiac valves.

The formation of myofibroblasts in valve interstitial cell (VIC) populations contributes to fibrotic valvular disease. We examined myofibroblast differentiation in VICs from porcine aortic valves. In normal valves, cells immunostained for alpha-smooth muscle actin (alpha-SMA, a myofibroblast marker) were rare (0.69 +/- 0.48%), but in sclerotic valves of animals fed an atherogenic diet, myofibroblasts were spatially clustered and abundant (31.2 +/- 6.3%). In cultured VIC populations from normal valves, SMA-positive myofibroblasts were also spatially clustered, abundant (21% positive cells after 1 passage), and stained for collagen type I and vimentin but not desmin. For an analysis of stem cells, two-color flow cytometry of isolated cells stained with Hoechst 33342 demonstrated that 0.5% of VICs were side population cells; none stained for SMA. Upon culture, sorted side population cells generated approximately 85% SMA-positive cells, indicating that some myofibroblasts originate from a rare population with stem cell characteristics. Plating cells on rigid collagen substrates enabled the formation of myofibroblasts after 5 days in culture, which was completely blocked by culture of cells on compliant collagen substrates. Exogenous tensile force also significantly increased SMA expression in VICs. Isotope-coded affinity tags and mass spectrometry were used to identify differentially expressed proteins in myofibroblast differentiation of VICs. Of the nine proteins that were identified, cofilin expression and phospho-cofilin were strongly increased by conditions favoring myofibroblast differentiation. Knockdown of cofilin with small-interfering RNA inhibited collagen gel contraction and reduced myofibroblast differentiation as assessed by the SMA incorporation into stress fibers. When compared with normal valves, diseased valves showed strong immunostaining for cofilin that colocalized with SMA in clustered cells. We conclude that in VICs, cofilin is a marker for myofibroblasts in vivo and in vitro that arise from a rare population of stem cells and require a rigid matrix for formation.

Biliary duct ablation with N-butyl cyanoacrylate.

PURPOSE: To assess the efficacy of percutaneous insertion of n-butyl cyanoacrylate (NBCA) in the ablation of bile ducts in patients with persistent postsurgical bile leaks in which traditional means of treatment have failed. MATERIALS AND METHODS: Ablation of bile ducts with NBCA was performed in six patients (two men and four women). The average length of follow-up was 27 months (range, 13-46 months). Four patients presented after hepatic lobectomy with a persistent bile leak, one patient presented after cholecystectomy with a chronically obstructed bile duct, and one patient presented after cholecystectomy from intraoperative bile duct injury. After access to the biliary system was obtained, a cholangiogram was obtained. After the desired duct was isolated, it was copiously irrigated with saline solution. A glue solution containing NBCA glue, Ethiodol, and tantalum powder was delivered into the duct through a polyethylene catheter that had been irrigated with dextrose solution. RESULTS: Four patients had problems arising from isolated segmental ductal systems that had no communication with the normal biliary ductal system and were treated successfully on the first attempt. In two patients, there was communication to the main biliary ductal system and a persistent bile leak occurred that required placement of a coil and a second final gluing procedure. The only complication observed was unintentional spillage of glue into the main biliary system in one patient, which was ultimately clinically insignificant. CONCLUSIONS: The use of NBCA glue in obliteration of bile ducts is a safe procedure with excellent results in patients with complications from isolated segmental ducts. Although a repeat procedure may be necessary if the duct communicates with the main biliary tree, the procedure can decrease the morbidity associated with chronic external biliary drainage.

In situ synthesis of oligonucleotide arrays by using surface tension.

This work describes the in situ synthesis of oligonucleotide arrays on glass surfaces. These arrays are composed of features defined and separated by differential surface tension (surface tension arrays). Specifically, photolithographic methods were used to create a series of spatially addressable, circular features containing an amino-terminated organosilane coupled to the glass through a siloxane linkage. Each feature is bounded by a perfluorosilanated surface. The differences in surface energies between the features and surrounding zones allow for chemical reactions to be readily localized within a defined site. The aminosilanation process was analyzed using contact angle, X-ray photoelectron spectroscopy (XPS), and time-of-flight/secondary ion mass spectroscopy (TOF-SIMS). The efficiency of phosphoramidite-based oligonucleotide synthesis on these surface tension arrays was measured by two methods. One method, termed step-yields-by-hybridization, indicates an average synthesis efficiency for all four (A,G,C,T) bases of 99.9 +/- 1.1%. Step yields measured for the individual amidite bases showed efficiencies of 98.8% (dT), 98.0% (dA), 97.0% (dC), and 97.6% (dG). The second method for determining the amidite coupling efficiencies was by capillary electrophoresis (CE) analysis. Homopolymers of dT (40- and 60mer), dA (40mer), and dC (40mer) were synthesized on an NH(4)OH labile linkage. After cleavage, the products were analyzed by CE. Synthesis efficiencies were calculated by comparison of the full-length product peak with the failure peaks. The calculated coupling efficiencies were 98.8% (dT), 96.8% (dA), and 96.7% (dC).

Utilization of new technologies in drug trials and discovery.

It has become widely accepted that individual genetic variation is a prime determinant in both disease susceptibility and toxic response to therapeutic agents and xenobiotics. Emerging genetic sequence data and phenotype association studies are expected to enable disease risk prediction and guide subsequent therapeutic approaches in individual cases. However, making a good match between an individual genetic profile, disease risk prediction, and appropriate therapeutic intervention will require genotyping many polymorphic sites in large numbers of genes or single nucleotide polymorphism sites throughout the genome. Additionally, each polymorphism will have to be associated with a phenotype. Presumably, a composite phenotype may be predicted by integrating anticipated contributions from each polymorphism contributing to the complex genotype. Methods for executing such large-scale genotyping studies are rapidly evolving and becoming available. DNA microarray technology applied in hybridization-based genotyping assays is particularly well suited to respond to the accelerating pace of polymorphism discovery and the associated demand for highly parallel genotyping capability.

Extension of a pilot study: impact from the cytochrome P450 2D6 polymorphism on outcome and costs associated with severe mental illness.

The influence of cytochrome P450 2D6 (CYP2D6) genetic variability was examined in psychiatric inpatients by evaluating adverse drug events (ADEs), hospital stays, and total costs over a 1-year period in an extension of a previously published brief report. One hundred consecutive psychiatric patients from Eastern State Hospital in Lexington, Kentucky, were genotyped for CYP2D6 expression. ADEs were evaluated by a neurologic rating scale, modified Udvalg for Kliniske Undersogelser Side Effect Rating Scale, or chart review. Information on total hospitalization days and total costs were gathered for a 1-year period. Forty-five percent of the patients received medications that were primarily dependent on the CYP2D6 enzyme for their elimination. When the analysis was restricted to just those patients in each group receiving medication heavily dependent on the CYP2D6 enzyme, the following were observed: (1) a trend toward greater numbers of ADEs from medications as one moved from the group with ultrarapid CYP2D6 activity (UM) to the group with absent CYP2D6 activity (PM); (2) the cost of treating patients with extremes in CYP2D6 activity (UM and PM) was on average $4,000 to $6,000 per year greater than the cost of treating patients in the efficient metabolizer (EM) and intermediate metabolizer (IM) groups; and (3) total duration of hospital stay was more pronounced for those in CYP2D6 PM group. Variance of hospital stays and costs calculated from these preliminary data suggests that 1,500 to 2,000 patients must be evaluated over at least a 1-year period to determine whether the CYP2D6 genetic variation significantly alters the duration of hospital stay and costs.

JC virus enters human glial cells by clathrin-dependent receptor-mediated endocytosis.

The human polyomavirus JC virus (JCV) is the etiologic agent of a fatal central nervous system (CNS) demyelinating disease known as progressive multifocal leukoencephalopathy (PML). PML occurs predominantly in immunosuppressed patients and has increased dramatically as a result of the AIDS pandemic. The major target cell of JCV infection and lytic replication in the CNS is the oligodendrocyte. The mechanisms by which JCV initiates and establishes infection of these glial cells are not understood. The initial interaction between JCV and glial cells involves virus binding to N-linked glycoproteins containing terminal alpha(2-6)-linked sialic acids. The subsequent steps of entry and targeting of the viral genome to the nucleus have not been described. In this report, we compare the kinetics and mechanisms of infectious entry of JCV into human glial cells with that of the related polyomavirus, simian virus 40 (SV40). We demonstrate that JCV, unlike SV40, enters glial cells by receptor-mediated clathrin-dependent endocytosis.