The cystine-glutamate transporter SLC7A11 has been implicated in chemoresistance, by supplying cystine to the cell for glutathione maintenance. In the NCI-60 cell panel, SLC7A11 expression shows ...negative correlation with growth inhibitory potency of geldanamycin but not with its analog 17-(allylamino)-17-demethoxygeldanamycin (17-AAG), which differs in the C-17 substituent in that the the methoxy moiety of geldanamycin is replaced by an amino group. Structure and potency analysis classified 18 geldanamycin analogs into two subgroups, "17-O/H" (C-17 methoxy or unsubstituted) and "17-N" (C-17 amino), showing distinct SLC7A11 correlation. We used three 17-O/H analogs and four 17-N analogs to test the role of the 17-substituents in susceptibility to SLC7A11-mediated resistance. In A549 cells, which are resistant to geldanamycin and strongly express SLC7A11, inhibition of SLC7A11 by (S)-4-carboxyphenylglycine or small interfering RNA increased sensitivity to 17-O/H, but had no effect on 17-N analogs. Ectopic expression of SLC7A11 in HepG2 cells, which are sensitive to geldanamycin and express low SLC7A11, confers resistance to geldanamycin, but not to 17-AAG. Antioxidant N-acetylcysteine, a precursor for glutathione synthesis, completely suppressed cytotoxic effects of 17-O/H but had no effect on 17-N analogs, whereas the prooxidant ascorbic acid had the opposite effect. Compared with 17-AAG, geldanamycin led to significantly more intracellular reactive oxygen species (ROS) production, which was quenched by addition of N-acetylcysteine. We conclude that SLC7A11 confers resistance selectively to 17-O/H (e.g., geldanamycin) but not to 17-N (e.g., 17-AAG) analogs partly as a result of differential dependence on ROS for cytotoxicity. Distinct mechanisms could significantly affect antitumor response and organ toxicity of these compounds in vivo.
Introduction: Unnatural amino acids (UAAs) share the same basic structure as proteinogenic amino acids. However, UAAs permit additional functions and applications to proteins due to their different ...side chains. Recent UAA applications include using fluorescent UAAs to label proteins. The UAA system provides an alternative method to traditional protein labeling mechanisms (antibodies, GFP, and tags, such as HA and HIS), which can affect protein functionality and topology. The purpose of this study was to visualize the hepatitis C virus (HCV) core protein using the fluorescent UAA Anap (3-(6-acetyl-2-naphthalenyl)amino-L-alanine). Methods: Huh-7.5 cells were co-transfected with HCV core plasmids containing amber stop codons at various positions throughout the coding sequence and a second plasmid encoding the orthogonal tRNA/synthetase pair that facilitates Anap incorporation. Three days post transfection, cells were stained for core protein and lipid droplets (LDs) and visualized using immunofluorescence or confocal microscopy. Results: We have optimized transfection protocols for the efficient expression of the tRNA/synthetase pair required for Anap incorporation and are able to visualize our core mutant proteins containing Anap. We have successfully substituted Anap into 11 different positions within the core, including substitutions for tryptophan, tyrosine, and phenylalanine residues. In addition, we have shown that our core mutants associate with cellular LDs, suggesting that the incorporation of the UAA did not disrupt core protein expression, stability, or cellular localization. Conclusions: We have demonstrated the establishment of a UAA incorporation system in an HCV protein without any obvious impact on core protein function. The ability to label viral proteins using fluorescent UAAs eliminates the requirement of antibodies or tags for protein visualization. In conclusion, the UAA system is a useful method to study HCV proteins and can potentially be used to label viruses for live cell and animal studies.
Introduction
Severe traumatic brain injury (TBI) is the world’s leading cause of permanent neurological disability in children. TBI-induced neurological deficits may be driven by neuroinflammation ...post-injury. Abnormal activity of SH2 domain-containing inositol 5′ phosphatase-1 (SHIP-1) has been associated with dysregulated immunological responses, but the role of SHIP-1 in the brain remains unclear. The current study investigated the immunoregulatory role of SHIP-1 in a mouse model of moderate–severe pediatric TBI.
Methods
SHIP-1+/− and SHIP-1−/− mice underwent experimental TBI or sham surgery at post-natal day 21. Brain gene expression was examined across a time course, and immunofluorescence staining was evaluated to determine cellular immune responses, alongside peripheral serum cytokine levels by immunoassays. Brain tissue volume loss was measured using volumetric analysis, and behavior changes both acutely and chronically post-injury.
Results
Acutely, inflammatory gene expression was elevated in the injured cortex alongside increased IBA-1 expression and altered microglial morphology; but to a similar extent in SHIP-1−/− mice and littermate SHIP-1+/− control mice. Similarly, the infiltration and activation of CD68-positive macrophages, and reactivity of GFAP-positive astrocytes, was increased after TBI but comparable between genotypes. TBI increased anxiety-like behavior acutely, whereas SHIP-1 deficiency alone reduced general locomotor activity. Chronically, at 12-weeks post-TBI, SHIP-1−/− mice exhibited reduced body weight and increased circulating cytokines. Pro-inflammatory gene expression in the injured hippocampus was also elevated in SHIP-1−/− mice; however, GFAP immunoreactivity at the injury site in TBI mice was lower. TBI induced a comparable loss of cortical and hippocampal tissue in both genotypes, while SHIP-1−/− mice showed reduced general activity and impaired working memory, independent of TBI.
Conclusion
Together, evidence does not support SHIP-1 as an essential regulator of brain microglial morphology, brain immune responses, or the extent of tissue damage after moderate–severe pediatric TBI in mice. However, our data suggest that reduced SHIP-1 activity induces a greater inflammatory response in the hippocampus chronically post-TBI, warranting further investigation.
To characterize the incidence of and risk factors for a detectable drug level (DDL) in patients that received inhaled aminoglycoside therapy.
This retrospective, single-centre study included adult ...patients who received at least one dose of an inhaled aminoglycoside with a drug level during inpatient hospitalization. Patients were excluded if they received an aminoglycoside intravenously within 7 days or if the drug level was not drawn within 4 h of the next dose. A repeated measures logistic regression model evaluated the association between potential risk factors and a DDL.
Among 286 drug levels, 88 (30.8%) drug levels were detectable. In multivariable analysis, cystic fibrosis (CF) (OR: 3.03; 95% CI: 1.10-8.35), chronic kidney disease (CKD) (OR: 4.25; 95% CI: 1.84-9.83), lung transplant recipient (OR: 3.08; 95% CI: 1.09-8.73), mechanical ventilation (OR: 2.99; 95% CI: 1.25-7.15) and tobramycin (OR: 5.26; 95% CI: 2.35-11.78) were associated with higher odds of a DDL. Among those with a DDL, inhaled aminoglycoside type and drug level concentration were not associated with acute kidney injury (P = 0.161).
Among 286 drug levels identified among inpatients receiving inhaled aminoglycoside therapy, 88 (30.8%) unique drug levels were detectable. Based on the results of this study, periodic trough concentrations should be considered for patients receiving inhaled aminoglycoside therapy with CF, CKD, lung transplantation, mechanical ventilation or tobramycin.
Posttraumatic arthritis (PTA) occurs commonly after articular fracture and may arise, in part, from joint surface incongruity after injury. MRL/MpJ (MRL) “super‐healer” mice are protected from PTA ...compared to C57BL/6 (B6) mice following articular fracture. However, the relationship between the initial displacement of the articular surface, biologic response, and susceptibility to PTA after fracture remains unclear. The objective of this study was to assess whether joint incongruity after articular fracture, as measured by in vivo micro‐computed tomography (microCT), could predict pathomechanisms of PTA in mice. B6 and MRL mice (n = 12/strain) received a closed articular fracture (fx) of the left tibial plateau. Articular incongruity was quantified as bone surface deviations (BSD) for each in vivo microCT scan obtained from pre‐fx to 8 weeks post‐fx, followed by histologic assessment of arthritis. Serum concentrations of bone formation (PINP) and bone resorption (CTX‐I) biomarkers were quantified longitudinally. Both strains showed increases in surface incongruity over time, as measured by increases in BSD. In B6 mice, acute surface incongruity was significantly correlated to the severity of PTA (R
2 = 0.988; p = .0006), but not in MRL mice (R
2 = 0.224; p = .220). PINP concentrations significantly decreased immediately post‐fx in B6 mice (p = .023) but not in MRL mice, indicating higher bone synthesis in MRL mice. MRL/MpJ mice demonstrate a unique biologic response to articular fracture such that the observed articular bone surface displacement does not correlate with the severity of subsequent PTA. Clinical Relevance: Identifying therapies to enhance acute biologic repair following articular fracture may mitigate the risk of articular surface displacement for PTA.
Codeine: Time to Say "No" Tobias, Joseph D; Green, Thomas P; Coté, Charles J
Pediatrics,
10/2016, Letnik:
138, Številka:
4
Journal Article
Recenzirano
Odprti dostop
Codeine has been prescribed to pediatric patients for many decades as both an analgesic and an antitussive agent. Codeine is a prodrug with little inherent pharmacologic activity and must be ...metabolized in the liver into morphine, which is responsible for codeine's analgesic effects. However, there is substantial genetic variability in the activity of the responsible hepatic enzyme, CYP2D6, and, as a consequence, individual patient response to codeine varies from no effect to high sensitivity. Drug surveillance has documented the occurrence of unanticipated respiratory depression and death after receiving codeine in children, many of whom have been shown to be ultrarapid metabolizers. Patients with documented or suspected obstructive sleep apnea appear to be at particular risk because of opioid sensitivity, compounding the danger among rapid metabolizers in this group. Recently, various organizations and regulatory bodies, including the World Health Organization, the US Food and Drug Administration, and the European Medicines Agency, have promulgated stern warnings regarding the occurrence of adverse effects of codeine in children. These and other groups have or are considering a declaration of a contraindication for the use of codeine for children as either an analgesic or an antitussive. Additional clinical research must extend the understanding of the risks and benefits of both opioid and nonopioid alternatives for orally administered, effective agents for acute and chronic pain.
Community Empowerment Partners (CEPs) Hempstead, Bridgette; Green, Cynthia; Briant, Katherine J. ...
Journal of community health,
10/2018, Letnik:
43, Številka:
5
Journal Article
Recenzirano
Odprti dostop
Peer educators have been shown to provide effective interventions in breast cancer screening. Few studies have compared the effects of peer education on breast cancer knowledge among peer educators ...and the community members who are subsequently reached through the peer education. Further, little is known as to whether those who received the education then go on to educate others in the community. The purpose of this study is to address those gaps. Using a pre- and post-test study design, we trained peer educators, provided the educators with resources to train community members, and assessed changes in knowledge. We sought to train ten educators and recommended each train ten community members in breast cancer knowledge and screening strategies. A total of 14 peer educators were trained, who subsequently trained a total of 121 community members, of whom 94 were African American women. Peer educators and community members, showed comparable increases in knowledge. Community members who were educated also increased intention to discuss breast cancer and breast cancer screening with their family, friends, and acquaintances. Our study suggests that it is feasible to train peer educators to increase knowledge among community members to the same level that they themselves experience when trained. Further, community members are interested in sharing information learned related to how much they learn from peer educators.
Medication administration errors that take place in the home are common, especially when liquid preparations are used and complex medication schedules with multiple medications are involved; children ...with chronic conditions are disproportionately affected. Parents and other caregivers with low health literacy and/or limited English proficiency are at higher risk for making errors in administering medications to children in their care. Recommended strategies to reduce home medication errors relate to provider prescribing practices; health literacy-informed verbal counseling strategies (eg, teachback and showback) and written patient education materials (eg, pictographic information) for patients and/or caregivers across settings (inpatient, outpatient, emergency care, pharmacy); dosing-tool provision for liquid medication measurement; review of medication lists with patients and/or caregivers (medication reconciliation) that includes prescription and over-the-counter medications, as well as vitamins and supplements; leveraging the medical home; engaging adolescents and their adult caregivers; training of providers; safe disposal of medications; regulations related to medication dosing tools, labeling, packaging, and informational materials; use of electronic health records and other technologies; and research to identify novel ways to support safe home medication administration.
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