Post-Transcriptional Regulation of mRNA Metabolism during Differentiation of 3T3-L1 Cells: Role of HuR

Our studies address early control points in the differentiation process that are necessary for expression as well as the maintenance of the adipocyte phenotype. We have focused on the post-transcriptional regulation of mRNA metabolism by the RNA binding protein HuR. Depending on the particular mRNA, HuR has been suggested to control polyadenylation, translocation to the cytosol, mRNA stability, and/or translational efficiency. Using the 3T3-L1 preadipocyte cell line, we have demonstrated a critical role for HuR in the differentiation process by use of siRNA mediated suppression which resulted in an inhibition of differentiation. We have identified several mRNAs that serve as HuR ligands early in the differentiation process, one of which is C/EBPβ, an important adipocyte transcription factor, whose expression is highly controlled and essential for proper acquisition of the adipocyte phenotype. Within minutes of induction of differentiation, HuR forms an mRNP complex with C/EBPβ mRNA in the nucleus followed by translocation to the cytoplasm. HuR remains associated with C/EBPβ mRNA suggesting a role in mRNA stability and translation efficiency. Our data suggest that formation of this complex in the nucleus serves as a regulator/attenuator of polyadenylation and that this interaction leads to a controlled metabolism of the C/EBPβ mRNA by determination of the quantity of message translocated to the cytosol and available for translation. Additionally, our data have directed us toward the Zfp206 mRNA as an important HuR ligand whose regulation is predicted to be responsible for maintenance of the differentiation potential of the cells and whose expression is terminated as the cells express PPARγ and establish the adipocyte phenotype. Our mechanistic analysis of these issues will identify novel control points in the initial stages of adipogenesis and thus fundamental to the pathological states of obesity and diabetes.  Ph.D

HuR involvement in mitotic clonal expansion during acquisition of the adipocyte phenotype

In the nucleus HuR binds to mRNAs containing adenylate-uridylate rich elements in the 3?- untranslated region. HuR may influence expression of its ligand mRNA through regulation of polyadenylation, translocation of the message to the cytosol, stabilization of the mRNA and/or altering its translational efficiency. Suppression of HuR using siRNA resulted in an attenuation of the 3T3-L1 differentiation program, consistent with HuR control of the expression of mRNA ligand (s) critical to the differentiation process. In the current study we begin to identify mRNA ligands of HuR whose regulated expression is necessary for adipogenesis. Originally published in Biochemical and Biophysiological Research Communications Vol. 383, No. 2 2009

Topical and Systemic Cannabidiol Improves Trinitrobenzene Sulfonic Acid Colitis in Mice

Background/Aims: Compounds of Cannabis sativa are known to exert anti-inflammatory properties, some of them without inducing psychotropic side effects. Cannabidiol (CBD) is such a side effect-free phytocannabinoid that improves chemically induced colitis in rodents when given intraperitoneally. Here, we tested the possibility whether rectal and oral application of CBD would also ameliorate colonic inflammation, as these routes of application may represent a more appropriate way for delivering drugs in human colitis. Methods: Colitis was induced in CD1 mice by trinitrobenzene sulfonic acid. Individual groups were either treated with CBD intraperitoneally (10 mg/kg), orally (20 mg/kg) or intrarectally (20 mg/kg). Colitis was evaluated by macroscopic scoring, histopathology and the myeloperoxidase (MPO) assay. Results: Intraperitoneal treatment of mice with CBD led to improvement of colonic inflammation. Intrarectal treatment with CBD also led to a significant improvement of disease parameters and to a decrease in MPO activity while oral treatment, using the same dose as per rectum, had no ameliorating effect on colitis. Conclusion: The data of this study indicate that in addition to intraperitoneal application, intrarectal delivery of cannabinoids may represent a useful therapeutic administration route for the treatment of colonic inflammation. Copyright (C) 2012 S. Karger AG, Base

Species-specific susceptibility to cannabis-induced convulsions

Background and Purpose. Numerous claims are made for cannabis' therapeutic utility upon human seizures, but concerns persist about risks. A potential confounder is the presence of both 9-tetrahydrocannabinol ( 9-THC), variously reported to be pro- and anti-convulsant, and cannabidiol (CBD), widely confirmed as anticonvulsant. Therefore, we investigated effects of prolonged exposure to different 9-THC/CBD cannabis extracts on seizure activity and associated measures of endocannabinoid (eCB) system signalling. Experimental Approach. Cannabis extract effects on in vivo neurological and behavioural responses, and on bioanalyte levels, were measured in rats and dogs. Extract effects on seizure activity were measured using electroencephalography-telemetry in rats. eCB signalling was also investigated using radioligand binding in cannabis extract-treated rats, and treatment-naïve rat, mouse, chicken, dog and human tissue. Key Results. Prolonged exposure to cannabis extracts caused spontaneous, generalised seizures, subserved by epileptiform discharges in rats, but not dogs, and produced higher 9-THC, but lower 11-hydroxy-THC (11-OH-THC) and CBD, plasma concentrations in rats versus dogs. In the same rats, prolonged exposure to cannabis also impaired cannabinoid type 1 receptor (CB1R)-mediated signalling. Profiling CB1R expression, basal activity, extent of activation and sensitivity to 9-THC suggested interspecies differences in eCB signalling, being more pronounced in a species that exhibited cannabis extract-induced seizures (rat) than a species that did not (dog). Conclusion and Implications. Sustained cannabis extract treatment caused differential seizure, behavioural and bioanalyte levels between rats and dogs. Supporting radioligand binding data suggest species differences in eCB signalling. Interspecies variations may have important implications for predicting cannabis-induced convulsions from animal models

Distinct Neurobehavioural Effects of Cannabidiol in Transmembrane Domain Neuregulin 1 Mutant Mice

The cannabis constituent cannabidiol (CBD) possesses anxiolytic and antipsychotic properties. We have previously shown that transmembrane domain neuregulin 1 mutant (Nrg1 TM HET) mice display altered neurobehavioural responses to the main psychoactive constituent of cannabis, Δ9-tetrahydrocannabinol. Here we investigated whether Nrg1 TM HET mice respond differently to CBD and whether CBD reverses schizophrenia-related phenotypes expressed by these mice. Adult male Nrg1 TM HET and wild type-like littermates (WT) received vehicle or CBD (1, 50 or 100 mg/kg i.p.) for 21 days. During treatment and 48 h after withdrawal we measured behaviour, whole blood CBD concentrations and autoradiographic receptor binding. Nrg1 HET mice displayed locomotor hyperactivity, PPI deficits and reduced 5-HT2A receptor binding density in the substantia nigra, but these phenotypes were not reversed by CBD. However, long-term CBD (50 and 100 mg/kg) selectively enhanced social interaction in Nrg1 TM HET mice. Furthermore, acute CBD (100 mg/kg) selectively increased PPI in Nrg1 TM HET mice, although tolerance to this effect was manifest upon repeated CBD administration. Long-term CBD (50 mg/kg) also selectively increased GABAA receptor binding in the granular retrosplenial cortex in Nrg1 TM HET mice and reduced 5-HT2A binding in the substantia nigra in WT mice. Nrg1 appears necessary for CBD-induced anxiolysis since only WT mice developed decreased anxiety-related behaviour with repeated CBD treatment. Altered pharmacokinetics in mutant mice could not explain our findings since no genotype differences existed in CBD blood concentrations. Here we demonstrate that Nrg1 modulates acute and long-term neurobehavioural effects of CBD, which does not reverse the schizophrenia-relevant phenotypes

THCCOOH concentrations in whole blood: Are they useful in discriminating occasional from heavy smokers?

Some forensic and clinical circumstances require knowledge of the frequency of drug use. Care of the patient, administrative, and legal consequences will be different if the subject is a regular or an occasional cannabis smoker. To this end, 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (THCCOOH) has been proposed as a criterion to help to distinguish between these two groups of users. However, to date this indicator has not been adequately assessed under experimental conditions. We carried out a controlled administration study of smoked cannabis with a placebo. Cannabinoid levels were determined in whole blood using tandem mass spectrometry. Significantly high differences in THCCOOH concentrations were found between the two groups when measured during the screening visit, prior to the smoking session, and throughout the day of the experiment. Receiver operating characteristic (ROC) curves were determined and two threshold criteria were proposed in order to distinguish between these groups: a free THCCOOH concentration below 3 µg/L suggested an occasional consumption (≤ 1 joint/week) while a concentration higher than 40 µg/L corresponded to a heavy use (≥ 10 joints/month). These thresholds were tested and found to be consistent with previously published experimental data. The decision threshold of 40 µg/L could be a cut-off for possible disqualification for driving while under the influence of cannabis. A further medical assessment and follow-up would be necessary for the reissuing of a driving license once abstinence from cannabis has been demonstrated. A THCCOOH level below 3 µg/L would indicate that no medical assessment is required

A moving boundary problem with a nonequilibrium interfacial boundary condition

M.S.LD Koffma

Harold Karschner Jr. Interview for the Veterans\u27 Voices Project

Harold “Wayne” Karschner (DOB: August 1, 1948) enlisted in the United States Air Force in February 1972. Mr. Karschner served at Chʻing-chʻüan Kang Air Base in Taiwan. Mr. Karschner left the Air Force having achieved the rank of Sergeant.https://corescholar.libraries.wright.edu/veterans_voices/1189/thumbnail.jp