Use of a Physiologically Based Pharmacokinetic Model for Rats to Study the Influence of Body Fat Mass and Induction of CYP1A2 on the Pharmacokinetics of TCDD

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a highly lipophilic chemical that distributes into adipose tissue, especially at low doses. However, at high doses TCDD sequesters in liver because it induces cytochrome P450 1A2 (CYP1A2) that binds TCDD. A physiologically based pharmacokinetic (PBPK) model was developed that included an inducible elimination rate of TCDD in the Sprague-Dawley rat. Objectives of this work were to characterize the influence of induction of CYP1A2 and adipose tissue mass fraction on the terminal elimination half-life (t(1/2)) of TCDD using this PBPK model. When the model assumes a fixed elimination of TCDD, t(1/2) increases with dose, due to hepatic sequestration. Because experimental data indicate that the t(1/2) of TCDD decreases with dose, the model was modified to include an inducible elimination rate. The PBPK model was then used to compare the t(1/2) after an increase of adipose tissue mass fraction from 6.9 to 70%. The model suggests that at low exposures, increasing adipose tissue mass increases the terminal t(1/2). However, at higher exposures, as CYP1A2 is induced, the relationship between adipose tissue mass and t(1/2) reaches a plateau. This demonstrates that an inducible elimination rate is needed in a PBPK model in order to describe the pharmacokinetics of TCDD. At low exposures these models are more sensitive to parameters related to partitioning into adipose tissue

Non-Native R1 Substitution in the S4 Domain Uniquely Alters Kv4.3 Channel Gating

The S4 transmembrane domain in Shaker (Kv1) voltage-sensitive potassium channels has four basic residues (R1–R4) that are responsible for carrying the majority of gating charge. In Kv4 channels, however, R1 is replaced by a neutral valine at position 287. Among other differences, Kv4 channels display prominent closed state inactivation, a mechanism which is minimal in Shaker. To determine if the absence of R1 is responsible for important variation in gating characteristics between the two channel types, we introduced the V287R mutant into Kv4.3 and analyzed its effects on several voltage sensitive gating transitions. We found that the mutant increased the voltage sensitivity of steady-state activation and altered the kinetics of activation and deactivation processes. Although the kinetics of macroscopic inactivation were minimally affected, the characteristics of closed-state inactivation and recovery from open and closed inactivated states were significantly altered. The absence of R1 can only partially account for differences in the effective voltage sensitivity of gating between Shaker and Kv4.3. These results suggest that the S4 domain serves an important functional role in Kv4 channel activation and deactivation processes, and also those of closed-state inactivation and recovery

Multicolour-banding fluorescence in situ hybridisation (mbanding-FISH) to identify recurrent chromosomal alterations in breast tumour cell lines

Recurrent chromosome breakpoints in tumour cells may point to cancer genes, but not many have been molecularly characterised. We have used multicolour-banding fluorescence in situ hybridisation (mbanding-FISH) on breast tumour cell lines to identify regions of chromosome break created by inversions, duplications, insertions and translocations on chromosomes 1, 5, 8, 12 and 17. We delineate a total of 136 regions of break, some of them occurring with high frequency. We further describe two examples of dual-colour FISH characterisation of breakpoints, which target the 1p36 and 5p11–12 regions. Both breaks involve genes whose function is unknown to date. The mbanding-FISH strategy constitutes an efficient first step in the search for potential cancer genes

AUX1-mediated root hair auxin influx governs SCFTIR1/AFB-type Ca2+ signaling

Auxin is a key regulator of plant growth and development, but the causal relationship between hormone transport and root responses remains unresolved. Here we describe auxin uptake, together with early steps in signaling, in Arabidopsis root hairs. Using intracellular microelectrodes we show membrane depolarization, in response to IAA in a concentration- and pH-dependent manner. This depolarization is strongly impaired in aux1 mutants, indicating that AUX1 is the major transporter for auxin uptake in root hairs. Local intracellular auxin application triggers Ca2+ signals that propagate as long-distance waves between root cells and modulate their auxin responses. AUX1-mediated IAA transport, as well as IAA- triggered calcium signals, are blocked by treatment with the SCFTIR1/AFB - inhibitor auxinole. Further, they are strongly reduced in the tir1afb2afb3 and the cngc14 mutant. Our study reveals that the AUX1 transporter, the SCFTIR1/AFB receptor and the CNGC14 Ca2+ channel, mediate fast auxin signaling in roots

Effects of PPARs Agonists on Cardiac Metabolism in Littermate and Cardiomyocyte-Specific PPAR-γ –Knockout (CM-PGKO) Mice

Understanding the molecular regulatory mechanisms controlling for myocardial lipid metabolism is of critical importance for the development of new therapeutic strategies for heart diseases. The role of PPARγ and thiazolidinediones in regulation of myocardial lipid metabolism is controversial. The aim of our study was to assess the role of PPARγ on myocardial lipid metabolism and function and differentiate local/from systemic actions of PPARs agonists using cardiomyocyte-specific PPARγ –knockout (CM-PGKO) mice. To this aim, the effect of PPARγ, PPARγ/PPARα and PPARα agonists on cardiac function, intra-myocyte lipid accumulation and myocardial expression profile of genes and proteins, affecting lipid oxidation, uptake, synthesis, and storage (CD36, CPT1MIIA, AOX, FAS, SREBP1-c and ADPR) was evaluated in cardiomyocyte-specific PPARγ –knockout (CM-PGKO) and littermate control mice undergoing standard and high fat diet (HFD). At baseline, protein levels and mRNA expression of genes involved in lipid uptake, oxidation, synthesis, and accumulation of CM-PGKO mice were not significantly different from those of their littermate controls. At baseline, no difference in myocardial lipid content was found between CM-PGKO and littermate controls. In standard condition, pioglitazone and rosiglitazone do not affect myocardial metabolism while, fenofibrate treatment significantly increased CD36 and CPT1MIIA gene expression. In both CM-PGKO and control mice submitted to HFD, six weeks of treatment with rosiglitazone, fenofibrate and pioglitazone lowered myocardial lipid accumulation shifting myocardial substrate utilization towards greater contribution of glucose. In conclusion, at baseline, PPARγ does not play a crucial role in regulating cardiac metabolism in mice, probably due to its low myocardial expression. PPARs agonists, indirectly protect myocardium from lipotoxic damage likely reducing fatty acids delivery to the heart through the actions on adipose tissue. Nevertheless a direct non- PPARγ mediated mechanism of PPARγ agonist could not be ruled out

Root Suberin Forms an Extracellular Barrier That Affects Water Relations and Mineral Nutrition in Arabidopsis

Though central to our understanding of how roots perform their vital function of scavenging water and solutes from the soil, no direct genetic evidence currently exists to support the foundational model that suberin acts to form a chemical barrier limiting the extracellular, or apoplastic, transport of water and solutes in plant roots. Using the newly characterized enhanced suberin1 (esb1) mutant, we established a connection in Arabidopsis thaliana between suberin in the root and both water movement through the plant and solute accumulation in the shoot. Esb1 mutants, characterized by increased root suberin, were found to have reduced day time transpiration rates and increased water-use efficiency during their vegetative growth period. Furthermore, these changes in suberin and water transport were associated with decreases in the accumulation of Ca, Mn, and Zn and increases in the accumulation of Na, S, K, As, Se, and Mo in the shoot. Here, we present direct genetic evidence establishing that suberin in the roots plays a critical role in controlling both water and mineral ion uptake and transport to the leaves. The changes observed in the elemental accumulation in leaves are also interpreted as evidence that a significant component of the radial root transport of Ca, Mn, and Zn occurs in the apoplast

A three-experiment examination of iliotibial band strain characteristics during different conditions using musculoskeletal simulation.

PURPOSE: Iliotibial band syndrome (ITBS) is a common chronic pathology mediated via excessive Iliotibial band (ITB) strain. The purpose using a three-experiment approach is to provide insight into the differences in strain between different athletic movements, the incidence of ITBS in females, the efficacy of different prophylactic modalities for ITBS and also the kinematic parameters associated with ITB strain. METHODS: Experiment 1 examined male and female athletes performing run, 45° cut and one-legged hop movements, experiment 2 observed males and females, whilst running in five different orthotic conditions and experiment 3 examined males and females riding a cycle ergometer at 70, 80 and 90RPM whilst in prophylactic knee brace and no-brace conditions. In each experiment, kinematics were obtained using a motion capture system and ITB strain was measured using a musculoskeletal simulation approach. RESULTS: In experiment 1 ITB strain was greater in the run (male=3.87% & female=4.37%; P<0.001) and cut (male=3.12% & female=4.06%; P<0.001) movements compared to hop (male=0.87% & female=1.54%). Experiment 2 showed that females exhibited increased ITB strain (male=6.34% & female=8.91%; P<0.05) and ITB strain velocity (male=57.17%/s & female=77.41%/s; P<0.05) and also in females that ITB strain velocity was greater (P≤0.01) in lateral (80.22%/s) and no-orthotic (83.01%/s) conditions compared to medial (72.58%/s) and off the shelf orthoses (74.52%/s). The regression analyses across movements showed that ITB strain was predicted by sagittal and coronal plane mechanics at the hip (R2=0.15-0.30; P<0.05) and sagittal, coronal and transverse plane kinematics at the knee joint (R2=0.15-0.22; P<0.05). CONCLUSION: Further insight is provided into differences in ITB strain across functional athletic movements, the increased incidence of ITBS in females and the parameters linked most strongly with ITB strain during different movements is provided; whilst also highlighting the prophylactic efficacy of medial and off the shelf orthoses in female runners

Atorvastatin Therapy during the Peri-Infarct Period Attenuates Left Ventricular Dysfunction and Remodeling after Myocardial Infarction

Although statins impart a number of cardiovascular benefits, whether statin therapy during the peri-infarct period improves subsequent myocardial structure and function remains unclear. Thus, we evaluated the effects of atorvastatin on cardiac function, remodeling, fibrosis, and apoptosis after myocardial infarction (MI). Two groups of rats were subjected to permanent coronary occlusion. Group II (n = 14) received oral atorvastatin (10 mg/kg/d) daily for 3 wk before and 4 wk after MI, while group I (n = 12) received equivalent doses of vehicle. Infarct size (Masson's trichrome-stained sections) was similar in both groups. Compared with group I, echocardiographic left ventricular ejection fraction (LVEF) and fractional area change (FAC) were higher while LV end-diastolic volume (LVEDV) and LV end-systolic and end-diastolic diameters (LVESD and LVEDD) were lower in treated rats. Hemodynamically, atorvastatin-treated rats exhibited significantly higher dP/dtmax, end-systolic elastance (Ees), and preload recruitable stroke work (PRSW) and lower LV end-diastolic pressure (LVEDP). Morphometrically, infarct wall thickness was greater in treated rats. The improvement of LV function by atorvastatin was associated with a decrease in hydroxyproline content and in the number of apoptotic cardiomyocyte nuclei. We conclude that atorvastatin therapy during the peri-infarct period significantly improves LV function and limits adverse LV remodeling following MI independent of a reduction in infarct size. These salubrious effects may be due in part to a decrease in myocardial fibrosis and apoptosis