Role of brain IL-1beta on fatigue after exercise-induced muscle damage

Brain cytokines, induced by various inflammatory challenges, have been linked to sickness behaviors, including fatigue. However, the relationship between brain cytokines and fatigue after exercise is not well understood. Delayed recovery of running performance after muscle-damaging downhill running is associated with increased brain IL-1beta concentration compared with uphill running. However, there has been no systematic evaluation of the direct effect of brain IL-1beta on running performance after exercise-induced muscle damage. This study examined the specific role of brain IL-1beta on running performance (either treadmill or wheel running) after uphill and downhill running by manipulating brain IL-1beta activity via intracerebroventricular injection of either IL-1 receptor antagonist (ra; downhill runners) or IL-1beta (uphill runners). Male C57BL/6 mice were assigned to the following groups: uphill-saline, uphill-IL-1beta, downhill-saline, or downhill-IL-1ra. Mice initially ran on a motor-driven treadmill at 22 m/min and -14% or +14% grade for 150 min. After the run, at 8 h (wheel cage) or 22 h (treadmill), uphill mice received intracerebroventricular injections of IL-1beta (900 pg in 2 microl saline) or saline (2 microl), whereas downhill runners received IL-1ra (1.8 microg in 2 microl saline) or saline (2 microl). Later (2 h), running performance was measured (wheel running activity and treadmill run to fatigue). Injection of IL-1beta significantly decreased wheel running activity in uphill runners (P<0.01), whereas IL-1ra improved wheel running in downhill runners (P<0.05). Similarly, IL-1beta decreased and Il-1ra increased run time to fatigue in the uphill and downhill runners, respectively (P<0.01). These results support the hypothesis that increased brain IL-1beta plays an important role in fatigue after muscle-damaging exercise.     

Profound bioenergetic abnormalities in peri-infarct myocardial regions

Regions of myocardial infarct (MI) are surrounded by a border zone (BZ) of normally perfused but dysfunctional myocardium. Although systolic dysfunction has been attributed to elevated wall stress in this region, there is evidence that intrinsic abnormalities of contractile performance exist in BZ myocardium. This study examined whether decreases of high-energy phosphates (HEP) and mitochondrial F(1)F(0)-ATPase (mtATPase) subunits typical of failing myocardium exist in BZ myocardium of compensated postinfarct remodeled hearts. Eight pigs were studied 6 wk after MI was produced by ligation of the left anterior descending coronary artery (LAD) distal to the second diagonal. Animals developed compensated LV remodeling with a decrease of ejection fraction from 54.6 +/- 5.4% to 31 +/- 2.1% (MRI) 5 wk after LAD occlusion. The remote zone (RZ) myocardium demonstrated modest decreases of ATP and mtATPase components. In contrast, BZ myocardium demonstrated profound abnormalities with ATP levels decreased to 42% of normal, and phosphocreatine-to-ATP ratio ((31)P-magnetic resonance spectroscopy) decreased from 2.06 +/- 0.19 in normal hearts to 1.07 +/- 0.10, with decreases in alpha-, beta-, OSCP, and IF(1) subunits of mtATPase, especially in the subendocardium. The reduction of myocardial creatine kinase isoform protein expression was also more severe in the BZ relative to the RZ myocardium. These abnormalities were independent of a change in mitochondrial content because the mitochondrial citrate synthase protein level was not different between the BZ and RZ. This regional heterogeneity of ATP content and expression of key enzymes in ATP production suggests that energetic insufficiency in the peri-infarct region may contribute to the transition from compensated LV remodeling to congestive heart failure.     

Kinetic and thermodynamic properties of an immobilized glucoamylase from a mesophilic fungus, Arachniotus citrinus

Purified glucoamylase from Arachniotus citrinus was immobilized on polyacrylamide gel with 70% yield of immobilization. The immobilization improved the pH optima, temperature optima, values of K(m), V(max), and activation energy. Irreversible thermal denaturation studies of soluble and immobilized glucoamylase indicated that immobilization decreased the entropy and enthalpy of deactivation by magnitudes and made the immobilized glucoamylase thermodynamically more stable.     

Array synthesis of progesterone receptor antagonists: 3-aryl-1,2-diazepines

New non-steroidal chemotypes are required for the development of drugs targeting the steroid hormone receptors. The parallel array synthesis of 3-aryl-1,2-diazepines employing solid-supported reagents is described. The resulting compounds demonstrated high affinity binding to the progesterone receptor.     

Frequency of cystathionine β-synthase 844INS68 polymorphism in Southern Iran

Iranian population with an Indo-European origin is one of the oldest populations in the world. Historical evidence suggests the close similarity in the origin of Iranian, European and north Indian population. However, there are few anthropological and genetic evidences on this subject. This study, which is the first report from Iran, was performed to investigate the genetic origin of Iranian population using a polymorphism in Cystathionine beta synthase (CBS) gene known as 844INS68bp in this respect, genomic DNA was extracted from the whole blood of 480 healthy normal blood donors referred to Fars Blood Transfusion Center, using a salting out method. The fragment containing 844INS68bp was amplified, the normal fragment was 174 bp and the fragment containing the insertion was 242 bp in length. Results indicated that 418 (87.08%) out of 480 individuals had a normal (N/N) genotype, 59 (12.29%) individuals were heterozygote (N/I) and 3 (0.63%) had homozygote a mutated genotype (I/I). The total frequency of 844INS68bp allele was found 6.8% which is similar to with the reported in White Caucasians. Comparison of the genotype of this study with the polymorphism in other populations revealed that Southern Iranian population has a great similarity with other Caucasians populations’ especially South Italy and North America while differed from East Asian and African populations. These results are in agreement with the result of other studied polymorphisms. Therefore, despite the great admixture of Iranian population with the neighboring non-Caucasian populations during the time, Iranian population still share a genetic background with other Caucasian populations.     

Genetic relationships among Pistacia species using AFLP markers

This study addresses the phylogenetic relationship between Pistacia species by amplified fragment length polymorphism (AFLP). The plant materials of this study consisted of a total of 44 accessions belonging to P. vera, P. eurycarpa, P. khinjuk, all subspecies of P. atlantica (atlantica, mutica, kurdica and cabulica), three unknown genotypes and three accessions, proposed to be hybrid from P. eurycarpa × P. atlantica. The accessions were from Iran, Turkey, USA and Syria. Six AFLP primer combinations produced a total of 475 fragments, with average of 79.16 fragments per primer pair, of which, 336 bands were polymorphic. Unweighted pair group method based on arithmetic average (UPGMA) analysis was performed on jaccard’s similarity coefficient matrix and also average similarity of each species. According to the results, two main clusters were developed and P. vera, P. eurycarpa, P. atlantica (subsp. atlantica, kurdica, mutica, cabulica) and the hybrid genotypes located in the first main cluster. P. khinjuk accessions from Iran and USA localized in second main cluster. The hybrid accessions located between eurycarpa and atlantica species and their hybrid nature between these two species were confirmed. One of the unknown accessions clustered with the hybrid ones and the two other were grouped closely with P. Khinjuk. According to this study, the closest species to P. vera was Eurycarpa group, followed by P. atlantica. UPGMA analysis separated P. atlantica subsp. mutica and cabulica from P. atlantica and P. eurycarpa. Subspecies mutica and cabulica were two closest genotypes; hence, P. atlantica subsp. mutica could be classified as a distinct species as P. mutica and the cabulica as a subspecies of P. mutica. This study revealed that P. eurycarpa is synonym for P. atlantica subsp. kurdica and should be considered distinct from P. atlantica; however, P. atlantica showed a closer genetic similarity to P. eurycarpa than the other species.     

The endangered cyprinodont <Emphasis Type="Italic">Aphanius ginaonis</Emphasis> (Holly, 1929) from southern Iran is a valid species: evidence from otolith morphology

Aphanius Nardo, 1827 (Actinopterygii, Cyprinodontidae) is a widely distributed genus in the Mediterranean and Persian Gulf area and includes several endangered species. The otolith morphology in Aphanius is known to represent a valuable tool for the taxonomy, and is also indicative for the genetic diversity of a particular population. The present study focuses on the otoliths of the endangered A. ginaonis (Holly, 1929), which is endemic to the Geno hot spring in southern Iran. The taxonomic status of A. ginaonis has repeatedly been questioned, and some scholars have argued that it merely represents a morphological variation of the widespread A. dispar. We present a comparison of the otolith morphology of A. ginaonis (52 specimens) with that of A. dispar (Rüppell, 1828) from the Mehran River Basin (southern Iran) (17 specimens) and an A. dispar population from the Persian Gulf coast of the United Arab Emirates (32 specimens). Our data obtained from SEM pictures, otolith morphometry and statistical analyses suggest that A. ginaonis represents a valid species. In A. ginaonis individuals with a standard length exceeding 23 mm, the otolith variables length–height and rostrum length represent useful complementary diagnostic characters discriminating this species from other Aphanius species. Besides ontogenetic variation, we found extremely high otolith form variability in A. ginaonis, including some otoliths with a morphology distinctly deviating from the basic morphology type. We hypothesize that these variations may be a result of the artificial introduction of A. dispar into the Geno hot spring during the last years and subsequent hybridisation.     

Synthesis and biological evaluation of fused thio- and selenopyrans as new indolocarbazole analogues with aryl hydrocarbon receptor affinity

A series of thio- and selenopyrans having two fused indole units, structurally related to indolocarbazoles, have been prepared and evaluated for aryl hydrocarbon receptor (AhR) affinity, leading to the identification of several new significant AhR ligands. In particular, the parent thiopyrano[2,3-b:6,5-b′]diindole and its derivative having a methyl group in the central ring, as well as the two corresponding selenopyrans, displayed the highest potencies of the compounds tested.     

Design,synthesis and SAR studies of 4-allyoxyaniline amides as potent 15-lipoxygensae inhibitors

A group of 4-allyloxyaniline amides 5a–o were designed, synthesized and evaluated as potential inhibitors of soybean 15-lipoxygenase (SLO) on the basis of eugenol and esteragol structures. Compound 5e showed the best IC₅₀ in SLO inhibition (IC₅₀ = 0.67 ± 0.06 μM). All compounds were docked in SLO active site retrieved from RCSB Protein Data Bank (PDB entry: 1IK3) and showed that allyloxy group of compounds is oriented towards the Fe³⁺-OH moiety in the active site of enzyme and fixed by hydrogen bonding with two conserved His⁵¹³ and Gln⁷¹⁶. It is resulted that molecular volume of the amide moiety would be a major factor in inhibitory potency variation of the synthetic amides, where the hydrogen bonding of the amide group could also involve in the activity of the inhibitors.     

EHD4 and CDH23 Are Interacting Partners in Cochlear Hair Cells

Cadherin 23 (CDH23), a transmembrane protein localized near the tips of hair cell stereocilia in the mammalian inner ear, is important for delivering mechanical signals to the mechano-electric transducer channels. To identify CDH23-interacting proteins, a membrane-based yeast two-hybrid screen of an outer hair cell (OHC) cDNA library was performed. EHD4, a member of the C-terminal EH domain containing a protein family involved in endocytic recycling, was identified as a potential interactor. To confirm the interaction, we first demonstrated the EHD4 mRNA expression in hair cells using in situ hybridization. Next, we showed that EHD4 co-localizes and co-immunoprecipitates with CDH23 in mammalian cells. Interestingly, the co-immunoprecipitation was found to be calcium-sensitive. To investigate the role of EHD4 in hearing, compound action potentials were measured in EHD4 knock-out (KO) mice. Although EHD4 KO mice have normal hearing sensitivity, analysis of mouse cochlear lysates revealed a 2-fold increase in EHD1, but no increase in EHD2 or EHD3, in EHD4 KO cochleae compared with wild type, suggesting that a compensatory increase in EHD1 levels may account for the absence of a hearing defect in EHD4 KO mice. Taken together, these data indicate that EHD4 is a novel CDH23-interacting protein that could regulate CDH23 trafficking/localization in a calcium-sensitive manner.Hair cells located in the mammalian inner ear transform mechanical stimuli into electrical signals that in turn facilitate neurotransmitter release onto auditory neurons. The key element in the transduction process is the mechano-electric transducer (MET)² apparatus located near the top of the stereocilium. CDH23 is a single pass transmembrane protein with 27 extracellular cadherin repeats. It is one of the components of the tip-link (1, 2), which connects the top of a shorter stereocilium to the side of its taller neighbor (3). Vibrations of the basilar membrane of the inner ear ultimately result in deflection of the hair bundles, which modulates tension on the tip-link, thereby controlling the opening probability of cation-selective MET channels (3, 4). Cations, principally K⁺ and Ca²⁺, flow through the MET channels and ultimately change the membrane potential. A mutation in the gene encoding CDH23, the Usher syndrome type 1D factor (USH1D), causes deaf-blindness in humans (5). Several interacting partners of CDH23 have been reported and include another tip-link protein protocadherin 15 (6), a multi-PDZ domain-containing scaffold protein harmonin (7) and a stereociliary scaffolding protein MAGI-1 (8). Protocadherin 15 binds to CDH23 through its extracellular domains (6), whereas the cytoplasmic region of CDH23 interacts with MAGI-1 and harmonin through its PDZ domain-binding interfaces (PBI). Harmonin also associates with other USH1 factors like myosin VIIa, protocadherin 15, and sans (9). These findings indicate that harmonin bridges CDH23 to the cytoskeletal actin core of the stereocilium and is probably essential for the developmental differentiation of stereocilia (10–12). However, it is currently unknown how CDH23 is transported to the tip of stereocilia. To search for additional interacting partners of CDH23, we performed a membrane-based yeast two-hybrid assay, which identified EHD4 as a potential binding partner (13).EHD4 belongs to an evolutionarily conserved EH (Eps 15 homology) domain-containing protein family involved in endocytic trafficking and recycling. Four highly homologous members of this family, EHD1–4, are expressed in mammalian cells. They contain a single C-terminal EH domain, an N-terminal nucleotide-binding loop and a coiled-coil region responsible for oligomerization (14–16). Of the four EHD proteins EHD1 is the best characterized and is involved in regulating the recycling of membrane receptors including the transferrin receptor and the major histocompatibility complex class I (17, 18). EHD1 is also involved in controlling cholesterol recycling and homeostasis (19) and in facilitating endosome to Golgi retrieval (20). EHD3 appears to regulate receptor movements from the early endosome (EE) to the endocytic recycling compartment (ERC) and Golgi (21, 22). EHD2 was isolated from GLUT4-enriched fractions of adipocytes and was shown to regulate insulin-mediated translocation of GLUT4 to the plasma membrane (23, 24). Additionally, EHD2 is involved in the regulation of transferrin receptor internalization (23), recycling (25), and actin cytoskeleton rearrangement (23). EHD4, also called Pincher, was first reported as an extracellular matrix protein (26). Subsequent studies have shown this intracellular protein to be involved in the regulation of neurotrophin receptor TrkA endocytosis in pheochromocytoma (PC12) cells (27). It is also involved in interactions with the cell fate determinant, NUMB, and co-localizes with the small GTP-binding protein, Arf6 (28). Recently, Sharma et al. (29) showed that EHD4 regulates the exit of endocytic cargo from the early endosome toward both the recycling compartment and the late endocytic pathway. They also indicated that EHD4 and EHD1 interact transiently as most of the EHD4 resides on peripheral early endosomes, while EHD1 resides primarily on tubular recycling compartments. This partial overlap/association might be necessary for the transport of proteins through the early endosome to the ERC. Previously, George et al. (25) had also demonstrated that EHD4 interacts with EHD1 and its paralogs, which suggests cooperation and partial overlap of function between EHD4 and EHD1.Unlike other CDH23-binding proteins, EHD4 does not contain a PDZ domain that could bind to the PBI located in the cytoplasmic tail of CDH23. In addition, the cytoplasmic tail of CDH23 lacks an Asn-Pro-Phe (NPF) motif that could mediate an interaction with the EH domain of EHD4. Therefore, we proceeded to characterize the authenticity of interaction between EHD4 and CDH23 identified in yeast and mammalian cells, using both in vitro and in vivo methods. We verified the expression of EHD4 mRNA in mouse cochlea and investigated the physiological role of EHD4 protein in the cochlea using EHD4-KO mice.