Pharmacogenetics: CYPs, NAT2 and 5-HTT Related to Antidepressants

Pharmacogenetics (PGt), the study of a gene\u27s influence on patient response to a drug, shows strong potential for explaining issues with efficacy related to antidepressant medications. Each year, antidepressants are one of the most commonly prescribed medications due to the millions of Americans affected by depression. Importantly, it is recognized that there is wide interpatient variability in drug response to antidepressants caused by genetic mutations, which can alter the pharmacodynamic (PD) and pharmacokinetic (PK) properties of various drugs used to treat depression. Proteins that are mainly involved in how patients respond to medications include receptors, drug-targeted proteins, drug transport proteins and drug-metabolizing enzymes. Specifically in depression, variations in the serotonin reuptake transporter (SERT-1 or 5-HTT), N-acetyltransferase (NAT2), cytochrome P450 (CYP) 2C19, 2D6, and 1A2 can affect the outcomes of patients receiving certain antidepressant medications. Utilizing PGt can help prevent the trial and error in prescribing antidepressants and lead to better patient outcomes in the treatment of depression. Pharmacists can utilize genetic information to help primary care physicians choose drug regimens that are more likely to benefit their patients. Although advances are being made in this subject matter, some major efforts of future research will evaluate the efficacy of drug regimens and the dosing of drugs based on patient genetics

Biological factors that impinge on Chagas disease drug development.

Chagas disease is caused by infection with the insect-transmitted protozoan Trypanosoma cruzi, and is the most important parasitic infection in Latin America. The current drugs, benznidazole and nifurtimox, are characterized by limited efficacy and toxic side-effects, and treatment failures are frequently observed. The urgent need for new therapeutic approaches is being met by a combined effort from the academic and commercial sectors, together with major input from not-for-profit drug development consortia. With the disappointing outcomes of recent clinical trials against chronic Chagas disease, it has become clear that an incomplete understanding of parasite biology and disease pathogenesis is impacting negatively on the development of more effective drugs. In addition, technical issues, including difficulties in establishing parasitological cure in both human patients and animal models, have greatly complicated the assessment of drug efficacy. Here, we outline the major questions that need to be addressed and discuss technical innovations that can be exploited to accelerate the drug development pipeline

Diverse voltage-sensitive dyes modulate GABAA receptor function

Voltage-sensitive dyes (VSDs) are important tools for assessing network and single-cell excitability, but an untested premise in most cases is that the dyes do not interfere with the parameters (membrane potential, excitability) that they are designed to measure. We found that popular members of several different families of voltage-sensitive dyes modulate GABA(A) receptor with maximum efficacy and potency similar to clinically used GABA(A) receptor modulators. Di-4-ANEPPS and DiBAC4(3) potentiated GABA function with micromolar and high nanomolar potency respectively and yielded strong maximum effects similar to barbiturates and neurosteroids. Newer blue oxonols had biphasic effects on GABA(A) receptor function at nanomolar and micromolar concentrations, with maximum potentiation comparable to that of saturating benzodiazepine effects. ANNINE 6 and ANNINE 6plus had no detectable effect on GABA(A) receptor function. Even dyes with no activity on GABA(A) receptors at baseline induced photodynamic enhancement of GABA(A) receptors. The basal effects of dyes were sufficient to prolong IPSCs and to dampen network activity in multielectrode array recordings. Therefore, the dual effects of voltage-sensitive dyes on GABAergic inhibition require caution in dye use for studies of excitability and network activity

Differential Reinforcement of Alternative Behavior Increases Resistance to Extinction: Clinical Demonstration, Animal Modeling, and Clinical Test of One Solution

Basic research with pigeons on behavioral momentum suggests that differential reinforcement of alternative behavior (DRA) can increase the resistance of target behavior to change. This finding suggests that clinical applications of DRA may inadvertently increase the persistence of target behavior even as it decreases its frequency. We conducted three coordinated experiments to test whether DRA has persistence-strengthening effects on clinically significant target behavior and then tested the effectiveness of a possible solution to this problem in both a nonhuman and clinical study. Experiment 1 compared resistance to extinction following baseline rates of reinforcement versus higher DRA rates of reinforcement in a clinical study. Resistance to extinction was substantially greater following DRA. Experiment 2 tested a rat model of a possible solution to this problem. Training an alternative response in a context without reinforcement of the target response circumvented the persistence-strengthening effects of DRA. Experiment 3 translated the rat model into a novel clinical application of DRA. Training an alternative response with DRA in a separate context resulted in lower resistance to extinction than employing DRA in the context correlated with reinforcement of target behavior. The value of coordinated bidirectional translational research is discusse

Intracellular DNA replication and differentiation of Trypanosoma cruzi is asynchronous within individual host cells in vivo at all stages of infection.

Investigations into intracellular replication and differentiation of Trypanosoma cruzi within the mammalian host have been restricted by limitations in our ability to detect parasitized cells throughout the course of infection. We have overcome this problem by generating genetically modified parasites that express a bioluminescent/fluorescent fusion protein. By combining in vivo imaging and confocal microscopy, this has enabled us to routinely visualise murine infections at the level of individual host cells. These studies reveal that intracellular parasite replication is an asynchronous process, irrespective of tissue location or disease stage. Furthermore, using TUNEL assays and EdU labelling, we demonstrate that within individual infected cells, replication of both mitochondrial (kDNA) and nuclear genomes is not co-ordinated within the parasite population, and that replicating amastigotes and non-replicating trypomastigotes can co-exist in the same cell. Finally, we report the presence of distinct non-canonical morphological forms of T. cruzi in the mammalian host. These appear to represent transitional forms in the amastigote to trypomastigote differentiation process. Therefore, the intracellular life-cycle of T. cruzi in vivo is more complex than previously realised, with potential implications for our understanding of disease pathogenesis, immune evasion and drug development. Dissecting the mechanisms involved will be an important experimental challenge

A novel role for thrombopoietin in regulating osteoclast development in humans and mice

Emerging data suggest that megakaryocytes (MKs) play a significant role in skeletal homeostasis. Indeed, osteosclerosis observed in several MK-related disorders may be a result of increased numbers of MKs. In support of this idea, we have previously demonstrated that MKs increase osteoblast (OB) proliferation by a direct cell-cell contact mechanism and that MKs also inhibit osteoclast (OC) formation. As MKs and OCs are derived from the same hematopoietic precursor, in these osteoclastogenesis studies we examined the role of the main MK growth factor, thrombopoietin (TPO) on OC formation and bone resorption. Here we show that TPO directly increases OC formation and differentiation in vitro. Specifically, we demonstrate the TPO receptor (c-mpl or CD110) is expressed on cells of the OC lineage, c-mpl is required for TPO to enhance OC formation in vitro, and TPO activates the mitogen-activated protein kinases, Janus kinase/signal transducer and activator of transcription, and nuclear factor-kappaB signaling pathways, but does not activate the PI3K/AKT pathway. Further, we found TPO enhances OC resorption in CD14+CD110+ human OC progenitors derived from peripheral blood mononuclear cells, and further separating OC progenitors based on CD110 expression enriches for mature OC development. The regulation of OCs by TPO highlights a novel therapeutic target for bone loss diseases and may be important to consider in the numerous hematologic disorders associated with alterations in TPO/c-mpl signaling as well as in patients suffering from bone disorders

Exploiting Genetically Modified Dual-Reporter Strains to Monitor Experimental Trypanosoma cruzi Infections and Host-Parasite Interactions.

Trypanosoma cruzi is the causative agent of Chagas disease, the most important parasitic infection in Latin America. Despite a global research effort, there have been no significant treatment advances for at least 40 years. Gaps in our knowledge of T. cruzi biology and pathogenesis have been major factors in limiting progress. In addition, the extremely low parasite burden during chronic infections has complicated the monitoring of both disease progression and drug efficacy, even in predictive animal models. To address these problems, we genetically modified T. cruzi to express a red-shifted luciferase. Mice infected with these highly bioluminescent parasites can be monitored by in vivo imaging, with exquisite sensitivity. However, a major drawback of bioluminescence imaging is that it does not allow visualization of host-parasite interactions at a cellular level. To facilitate this, we generated T. cruzi strains that express a chimeric protein that is both bioluminescent and fluorescent. Bioluminescence allows the tissue location of infection foci to be identified, and fluorescence can then be exploited to detect parasites in histological sections derived from excised tissue. In this article, we describe in detail the in vivo imaging and confocal microscopy protocols that we have developed for visualizing T. cruzi parasites expressing these dual-reporter fusion proteins. The approaches make it feasible to locate individual parasites within chronically infected murine tissues, to assess their replicative status, to resolve the nature of host cells, and to characterize their immunological context

In Vivo Analysis of Trypanosoma cruzi Persistence Foci at Single-Cell Resolution.

Infections with Trypanosoma cruzi are usually lifelong despite generating a strong adaptive immune response. Identifying the sites of parasite persistence is therefore crucial to understanding how T. cruzi avoids immune-mediated destruction. However, this is a major technical challenge, because the parasite burden during chronic infections is extremely low. Here, we describe an integrated approach involving comprehensive tissue processing, ex vivo imaging, and confocal microscopy, which allowed us to visualize infected host cells in murine tissue with exquisite sensitivity. Using bioluminescence-guided tissue sampling, with a detection level of 200 parasites, which we term mega-nests. In contrast, during the acute stage, when the total parasite burden is considerably higher and many cells are infected, nests containing >50 parasites are rarely found. In C3H/HeN mice, but not BALB/c mice, we identified skeletal muscle as a major site of persistence during the chronic stage, with most parasites being found in large mega-nests within the muscle fibers. Finally, we report that parasites are also frequently found in the skin during chronic murine infections, often in multiple infection foci. In addition to being a site of parasite persistence, this anatomical reservoir could play an important role in insect-mediated transmission and have implications for drug development.IMPORTANCE Trypanosoma cruzi causes Chagas disease, the most important parasitic infection in Latin America. Major pathologies include severe damage to the heart and digestive tract, although symptoms do not usually appear until decades after infection. Research has been hampered by the complex nature of the disease and technical difficulties in locating the extremely low number of parasites. Here, using highly sensitive imaging technology, we reveal the sites of parasite persistence during chronic-stage infections of experimental mice at single-cell resolution. We show that parasites are frequently located in smooth muscle cells in the circular muscle layer of the colon and that skeletal muscle cells and the skin can also be important reservoirs. This information provides a framework for investigating how the parasite is able to survive as a lifelong infection, despite a vigorous immune response. It also informs drug development strategies by identifying tissue sites that must be accessed to achieve a curative outcome

The Efficacy of Topical Hydrolyzed Psoralea corylifolia Extract in Treating Postinflammatory Hyperpigmentation

Background: Post-inflammatory hyperpigmentation (PIH) is common following resolution of acne. Purpose: The purpose of this study is to determine the treatment efficacy of Topical Hydrolyzed Psoralea Corylifolia extract (HPCE) on acne-induced PIH and TCA-induced PIH using a previously validated model.1Methods: A prospective, single-blinded, non-randomized study was conducted on 20 subjects with acne-induced PIH. Three acne-induced PIH areas on the face and three 35% TCA-induced PIH areas on the buttocks were analyzed. Subjects received topical HPCE [Unigen] and vehicle cream with instructions on twice daily application on two separate facial and gluteal lesions for 28 days; the third lesion served as a control. Clinical photography and Investigator Global Assessment (IGA) scores for hyperpigmentation were performed on days 0, 28, 35, 42, and 56 for all sites. Degree of improvement was defined as the change in the IGA score for hyperpigmentation between the first and last day of treatment. Results: For facial acne sites, one-way repeated measures ANOVA for degree of improvement as assessed by IGA analysis demonstrated a greater degree of improvement for product sites when compared to vehicle (1.9 times) and control (1.5 times); however, statistical significance was not reached. For TCA-induced PIH sites, there was a statistically significant degree of improvement for product treated sites compared to vehicle (9 times) and control (9 times). For both acne and TCA-induced PIH sites, Pearson correlation coefficient between time and IGA score for hyperpigmentation showed a strong and statistically significant (phttps://scholarlycommons.henryford.com/merf2019clinres/1010/thumbnail.jp

The reflective learning continuum: reflecting on reflection

The importance of reflection to marketing educators is increasingly recognized. However, there is a lack of empirical research which considers reflection within the context of both the marketing and general business education literature. This paper describes the use of an instrument which can be used to measure four identified levels of a reflection hierarchy: habitual action, understanding, reflection and intensive reflection and two conditions for reflection: instructor to student interaction and student to student interaction. Further we demonstrate the importance of reflective learning in predicting graduates’ perception of program quality. Although the focus was on assessment of MBA level curricula, the findings have great importance to marketing education and educators