Taurine transporter is expressed in vascular smooth muscle cells

Summary. The regulation of vascular smooth muscle cells (VSMCs) function by taurine has been a subject of increasing interest and investigation, and taurine is taken up into cells through a specific transporter system, the taurine transporter (TAUT). In the present study, we examined the expression of TAUT in VSMCs and the kinetic parameters of the uptake process of TAUT in VSMCs. RT-PCR and western blot demonstrated that the mRNA and protein of TAUT was expressed in VSMCs in vitro. Immunohistochemistry using antibody for TAUT revealed the expression of this protein in rat thoracic aorta. The maximal [³H]taurine uptake rate in VSMCs was 37.75 ± 3.13 pmol/min per mg of protein, with a K _m value of 5.42 ± 0.81 μM. Thus, VSMCs are able to express a functional taurine transporter. The regulation and detailed function of taurine and TAUT in VSMCs remain unclear, but our findings suggest a functional role for them in VSMCs metabolism.     

Freeze-fracture study of taste bud pores in the foliate papillae of the rabbit

Summary In freeze-fractured specimens of taste buds from the foliate papillae of rabbits, the intercellular spaces are separated from the pore of the taste bud by zonulae occludentes of the tight-type. Below these tight junctions numerous desmosomes are found at irregular intervals. The epithelial cells adjacent to the pore are also joined by single strands of fusion. The microvilli arising from the neck of the type I cells have a high particle density. The microvilli of type II cells and especially the short microvilli of peripherally situated cells have a lower intramembranous particle density. The single microvillus of type III cells has a very large diameter and is longer than the other microvilli. It contains a few larger intramembranous particles and vesicle-like protrusions of the membrane facing the cytoplasm. Transverse fracturing reveals a filamentous fine structure in all microvilli. The physiological implications of these observations are discussed.Supported by grants from the Deutsche Forschungsgemeinschaft (Ja 205/5+6)     

Developmental toxicity studies of lumefantrine and artemether in rats and rabbits

The combination of artemether plus lumefantrine is a type of artemisinin‐based combination therapy (ACT) recommended by the World Health Organization for uncomplicated falciparum malaria except in the first trimester of pregnancy. The first trimester restriction was based on the marked embryotoxicity in animals (including embryo death and cardiac and skeletal malformations) of artemisinins such as artesunate, dihydroartemisinin, and artemether. Before recommending ACTs for use in the first trimester, the World Health Organization has requested that all information relevant to the assessment of risk of ACTs to the embryo be made available to the public. This report describes the results of embryo‐fetal development studies of artemether alone, lumefantrine alone, and the combination in rats and rabbits as well as toxicokinetic studies of lumefantrine in pregnant rabbits. The developmental no‐effect levels for lumefantrine were 300 mg/kg/day in rats (based on a 25% decrease in litter size at 1000 mg/kg/day) and 1000 mg/kg/day in rabbits. The calculated safety margins based on human equivalent dose and plasma C_max and AUC values were in the range of 2.5‐ to 17‐fold. The developmental no‐effect levels for artemether were 3 mg/kg/day in rats and 25 mg/kg/day in rabbits. Lumefantrine caused no teratogenicity and was not a potent embryotoxin in rats and rabbits. Expected artemisinin‐like findings were seen with artemether alone and with artemether/lumefantrine combined except that no malformations were observed. There were no findings in pregnant rats and rabbits that would cause increased concern for the use of artemether–lumefantrine in the first trimester compared to other ACTs.     

The role of feeding,mating and ovariectomy on degeneration of indirect flight muscles of Dysdercus cingulatus (Heteroptera: Pyrrhocoridae)

Degeneration of indirect flight muscles takes place during the first gonotrophic cycle in females. Feeding and mating stimulate egg production and muscle histolysis. Starved virgin females do not histolyse the flight muscles. Mating has greater effect on muscle degeneration than feeding. Ovariectomy inhibits degeneration to a certain extent.     

Role of PCK2 in the proliferation of vascular smooth muscle cells in neointimal hyperplasia

Vascular smooth muscle cell (VSMC) proliferation is a hallmark of neointimal hyperplasia (NIH) in atherosclerosis and restenosis post-balloon angioplasty and stent insertion. Although numerous cytotoxic and cytostatic therapeutics have been developed to reduce NIH, it is improbable that a multifactorial disease can be successfully treated by focusing on a preconceived hypothesis. We, therefore, aimed to identify key molecules involved in NIH via a hypothesis-free approach. We analyzed four datasets ({"type":"entrez-geo","attrs":{"text":"GSE28829","term_id":"28829"}}GSE28829, {"type":"entrez-geo","attrs":{"text":"GSE43292","term_id":"43292"}}GSE43292, {"type":"entrez-geo","attrs":{"text":"GSE100927","term_id":"100927"}}GSE100927, and {"type":"entrez-geo","attrs":{"text":"GSE120521","term_id":"120521"}}GSE120521), evaluated differentially expressed genes (DEGs) in wire-injured femoral arteries of mice, and determined their association with VSMC proliferation in vitro. Moreover, we performed RNA sequencing on platelet-derived growth factor (PDGF)-stimulated human VSMCs (hVSMCs) post-phosphoenolpyruvate carboxykinase 2 (PCK2) knockdown and investigated pathways associated with PCK2. Finally, we assessed NIH formation in Pck2 knockout (KO) mice by wire injury and identified PCK2 expression in human femoral artery atheroma. Among six DEGs, only PCK2 and RGS1 showed identical expression patterns between wire-injured femoral arteries of mice and gene expression datasets. PDGF-induced VSMC proliferation was attenuated when hVSMCs were transfected with PCK2 siRNA. RNA sequencing of PCK2 siRNA-treated hVSMCs revealed the involvement of the Akt-FoxO-PCK2 pathway in VSMC proliferation via Akt2, Akt3, FoxO1, and FoxO3. Additionally, NIH was attenuated in the wire-injured femoral artery of Pck2-KO mice and PCK2 was expressed in human femoral atheroma. PCK2 regulates VSMC proliferation in response to vascular injury via the Akt-FoxO-PCK2 pathway. Targeting PCK2, a downstream signaling mediator of VSMC proliferation, may be a novel therapeutic approach to modulate VSMC proliferation in atherosclerosis.     

Deriving how far structural information is transmitted through parallel homodimeric coiled‐coils: A correlation analysis of helical staggers

How local conformation is affected by local sequence is fairly well understood for alpha‐helical coiled‐coils, but less is known about how local conformation is influenced by distant features. Here, I describe an approach to detect such an effect, based on computing correlation coefficients of local out‐of‐register alignments, or so‐called “staggers” between the helices, as a function of the axial distance between the staggers. This approach requires parallel homodimers, in which each stagger can occur with two “signs,” where either one helix or the other is shifted towards the N terminus. The signs of such staggers separated by up to 12 residues are strongly correlated, indicating that the conformations of the ends of coiled‐coils are commonly influenced by attached structures. Thus, the structures of coiled‐coil residues aberrantly attached to alternative proteins, such as those resulting from leukemogenic chromosomal rearrangements, may be distinguishable from those in normal tissues, and in turn serve as targets of selective drug design. The signs of helical staggers separated by between 13 and 30 residues are moderately yet significantly correlated, indicating that some of the coiled‐coils transmit this conformational feature axially for at least 45 Å. A positive, albeit noisy, correlation also exists among tropomyosin coiled‐coils for signed staggers separated by the 40‐residue actin repeat distance, consistent with the semi‐flexible tropomyosin filament binding F‐actin and regulating skeletal muscle contraction in a partially cooperative manner. Communication of the signs of axial staggers is explained in part by minimization of main‐chain hydrogen bond deformations. © Proteins 2013. © 2012 Wiley Periodicals, Inc.     

Ultrastructural changes in muscle mitochondria in situ,including the apparent development of internal septa,associated with the uptake and release of calcium

Treatment of mammalian muscle with the divalent cation ionophore A23187 causes the release of Ca2+ from the sarcoplasmic reticulum and allows the ultrastructural changes of the mitochondria during Ca2+-uptake to be demonstrated in situ. Electron micrographs reveal that the mitochondria swell dramatically during uptake, before contracting again when the accumulated Ca2+ is released once more into the cytoplasm. When maximally swollen, the mitochondria are apparently subdivided and internal "septa" are formed. The ultrastructural details concerning these internal membranous structures are shown in detail and their significance is discussed.     

Muscle palmitate uptake and binding are saturable and inhibited by antibodies to FABPPM

Studies show that uptake of long-chain fatty acids (LCFA) across the plasma membranes (PM) may occur partly via a carrier-mediated process and that the plasma membrane fatty acid-binding protein (FABP_PM) may be a component of this system. To test the hypothesis that FABP_PM is involved in transsarcolemmal transport of LCFA in muscle, we measured palmitate uptake in giant sarcolemmal vesicles and palmitate binding to PM proteins in rat muscles, (1) in the presence of increasing amounts of unbound palmitate and (2) in the absence or presence of antibody to FABP_PM. Both palmitate uptake and binding were found to be saturable functions of the unbound palmitate concentration with calculated V_max values of 10.5 ± 1.2 pmol/mg protein/15 sec and 45.6 ± 2.9 nmol/mg protein/15 min and K_m values of 12.8 ± 3.8 and 18.4 ± 1.8 nmol/L, respectively. The V_max values for both palmitate uptake and binding were significantly decreased by 75-79% in the presence of a polyclonal antibody to the rat hepatic FABP_PM. Antibody inhibition was found to be dose-dependent and specific to LCFA. Glucose uptake was not affected by the presence of the antibody to FABP_PM. Palmitate uptake and binding were also inhibited in the presence of trypsin and phloretin. These results support the hypothesis that transsarcolemmal LCFA transport occurs in part by a carrier-mediated process and that FABP_PM is a component of this process in muscle.     

Muscle ATP synthesis and utilisation,balanced during flow-induced increase of respiration

To investigate the control of cell energetic metabolism, creatine charge, ATP/ADP ratio and oxygen consumption (as indicators of an energetic status, the balance between ATP synthesis and degradation and the aerobic ATP turnover, respectively) were evaluated in the rat gracilis muscle, perfused-superfused in vitro. During the perfusion rate of 70 l/min the ATP/ADP ratio, as well as the creatine charge are kept at the in vivo level. With the decrease of the rate toward 54 l/min (of an abundant oxygen delivery), the values of both parameters are lower than levels in vivo. With the increase of the rate up to 100 l/min, both parameters are kept at the in vivo level, when respiration increases by 125%. The data demonstrate the 'unmatched' control of ATP utilisation and synthesis steady rates during the low perfusion rate; during the increasing steady ATP turnover following the increased perfusion rate, the two fluxes are strikingly 'matched', i.e. precisely balanced.