Is the early left‐right axis like a plant,a kidney,or a neuron? The integration of physiological signals in embryonic asymmetry

Embryonic morphogenesis occurs along three orthogonal axes. While the patterning of the anterior-posterior and dorsal-ventral axes has been increasingly well-characterized, the left-right (LR) axis has only relatively recently begun to be understood at the molecular level. The mechanisms that ensure invariant LR asymmetry of the heart, viscera, and brain involve fundamental aspects of cell biology, biophysics, and evolutionary biology, and are important not only for basic science but also for the biomedicine of a wide range of birth defects and human genetic syndromes. The LR axis links biomolecular chirality to embryonic development and ultimately to behavior and cognition, revealing feedback loops and conserved functional modules occurring as widely as plants and mammals. This review focuses on the unique and fascinating physiological aspects of LR patterning in a number of vertebrate and invertebrate species, discusses several profound mechanistic analogies between biological regulation in diverse systems (specifically proposing a nonciliary parallel between kidney cells and the LR axis based on subcellular regulation of ion transporter targeting), highlights the possible importance of early, highly-conserved intracellular events that are magnified to embryo-wide scales, and lays out the most important open questions about the function, evolutionary origin, and conservation of mechanisms underlying embryonic asymmetry.     

Comparative potency of some extended peptide chain members of the RFamide neuropeptide family,assessed on the hearts of<Emphasis Type="Italic"> Busycon canaliculatum</Emphasis> and<Emphasis Type="Italic"> Buccinum undatum</Emphasis>

Twenty different RFamide neuropeptide analogues were examined for their relative potencies on the ventricles of Busycon canaliculatum and Buccinum undatum and on the atrium of Busycon to determine the essential requirements for activity at the RFamide receptor. None of the neuropeptide studies was inhibitory to natural cardiac rhythmicity or to FMRFamide (Phe-Met-Arg-Phe-NH₂) or FLRFamide (Phe-Leu-Arg-Phe-NH₂) responses. Two tripeptides studied were completely without effect, indicating that a minimum of four amino acids in the peptide chain length was essential for any activity. The original parent tetrapeptide FMRFamide was surprisingly less potent than many of the extended chain peptides such as the penta, hepta and decapeptides. These RFamide neuropeptides were strongly inotropic on both ventricles and the atrium, while on the latter they were strongly chronotropic despite several of these peptides being of a non-molluscan origin. Chain length seems to be of little importance for activity at the receptor. Surprisingly, SCP_B (small cardioactive Peptide B) was not very effective in either Busycon or Buccinum ventricle. What was also clear was that the configuration of the carboxyl terminal was important for activity. Two neuropeptides in this study possessed an Arg-Met carboxyl terminal and were much less effective than FMRFamide, suggesting that an Arg-Phe terminal is most effective in receptor activation.Abbreviations Ala alanine - Arg arginine - Asn asparagine - Asp aspartic acid - FLRFamide Phe-Leu-Arg-Phe-NH₂ - FMRFamide Phe-Met-Arg-Phe-NH₂ - Glu glutamic acid - Gly glycine - His histidine - Leu leucine - LMS leucomyosuppressin - Met methionine - Nle norleucine - Phe phenylalanine - Pro proline - SCP_B Small cardioactive peptide B - Ser serine - Thr threonine - Trp tryptophan - Tyr tyrosine - Val valineCommunicated by G. HeldmaierPart of this work was presented at the Society for Experimental Biology 1999 Annual conference (Huddart et al. 1999)     

Hydrogen peroxide enhanced inron-induced injury in isolated heart and ventricular cardiomyocyte in rats

To explore the cardiac effects of iron with or without hydrogen peroxide, the isolated perfused rat heart and enzymatically isolated ventricular cardiomyocyte were used. It was shown that treatment with cell-permeable iron (Fe-HQ) for 10 min reduced the contractile amplitude and velocity and end diastolic cell length in the cardiomyocyte and increased the contents of lactate dehydrogenase (LDH) and creatine kinase (CK) in the coronary effluent and malondialdehyde (MDA) in the myocardium. The left ventricular developed pressure (LVDP), ± dP/dtmax, and heart rate and coronary flow are showed a biphasic phase, an increase at first followed by a decline. Treatment with hydrogen peroxide for 10 min following Fe-HQ augmented the effect of iron with an increase in coronary LDH and CK release and myocardial MDA content, and decrease in LVDP, ± dP/dtmax and heart rate. Perfusion of reduced glutathione with hydrogen peroxide counteracted these effects of Fe-HQ and hydrogen peroxide while dimethyl sulfoxide had no effect on the injury induced by Fe-HQ and hydrogen peroxide in the isolated rat heart. This suggests that augmentation of myocardial injury as a result of an increase in intracellular iron by hydrogen peroxide might involve the dysfunction of sulfydryl group containing proteins but not the hydroxyl radicals.     

Identification of nucleoside transport binding sites in the human myocardium

The role of nucleoside transport in ischemia-reperfusion injury and arrhythmias has been well documented in various animal models using selective blockers. However, clinical application of nucleoside transport inhibitors remains to be demonstrated in humans. It is not known whether human heart has nucleoside transport similar to that of animals. The aim of this study is to pharmacologically identify the presence of nucleoside transport binding sites in the human myocardium compared to animals.Myocardial tissue was obtained from guinea pig left and right ventricle, canine left ventricle, human intraoperative right atrium and human cadaveric right atrium and right and left ventricles. Myocardial preparations were obtained from tissue samples after homogenized and a differential centrifugation.Equilibrium binding assays were performed using [³H]-p-nitrobenzylthioinosine (NBMPR) at room temperature in the presence or absence of non-radioactive NBMPR or other nucleoside transport blockers such as p-nitrobenzylthioguanosine dipyridamole, lidoflazine, papaverin, adenosine and doxorubcine. From saturation curves and inhibition kinetics, we determined the relative maximal binding (B_max) and dissociation constant (K_d) of [³H]-NBMPR binding of human myocardial preparations.Results demonstrated that the fresh human myocardial preparations have a specific binding site for NBMPR with a B_max of 283 ± 32 fmol/mg protein and K_d of 0.56 ± 0.12 nM. These values are lower than those obtained from guinea pigs (B_max = 1440 ± 187 fmol/mg protein and K_d = 0.21 ± 0.03 nM) and canine atrium (B_max 594 ± 73 fmol/mg protein, and K_d = 1.12 ± 0.22 nM).Displacement kinetics studies revealed the relative potencies (of certain unrelated drugs as follow: p-nitrobenzylthioguanosine > dipyridamole > lidoflazine > pavaverine > Diltazam > adenosine > doxyrubicin. It is concluded that human myocardium contains an active nucleoside transport site which may play a crucial role in post-ischemic reperfusion-mediated injury in a wide spectrum of ischemic syndromes.     

Tissue and concentration-dependent effects of sodium selenite on muscle contraction

In this study, we demonstrated that sodium selenite with high doses (≥ 10^-3 M) were potent in inducing a contracture type effect on heart and smooth muscles. Selenite (Se), at a concentration of 10^-3 M, caused a contracture effect in heart preparations. Also, low Se concentrations did not have any significant effect. Although low concentrations of Se (≥ 10^-5 M) had a biphasic effects on acetylcholine (ACh) induced and spontaneous ileum contractions, 10^-3 M selenite enhanced once more a contracture effect similar to that of the heart preparations. Replacing Ca²⁺ concentration of the bathing solution by twofold Ca²⁺ or Ca²⁺-free did not change the effects of selenite (10^-5 M) on contractility of ileum preparations. In vascular smooth muscle, low concentration of selenite (<10^-4) had no significant effects on KC1, and phenylephrine-induced contractions and acethylcholineinduced endothelium-dependent relaxations of isolated rabbit aorta. However, the contractions induced by phenylephrine and the relaxations induced by acetylcholine in rabbit aorta were depressed significantly by high concentration of selenite (10^-3 M). The results obtained by selenite exposure from these three different types of tissue preparations first suggest that the high concentration of selenite exposure induces some alterations in the functions of muscles and endothelium in a tissue and dose-dependent manner. Second, this observed irreversible type of dysfunction of tissues induced by l0^-3 M selenite is not directly dependent on the Ca²⁺ entrance into the cytosol, but might be induced by the increase of intracellular Ca²⁺ with the disturbance of Ca²⁺ regulation.     

Initial hydrodynamic study on a new intraaortic axial flow pump: Dynamic aortic valve

Rotary blood pumps have been researched as implantable ventricular assist devices for years. To further reduce the complex of implanted axial pumps, the authors proposed a new concept of intraaortic axial pump, termed previously as “dynamic aortic valve (DAV)”. Instead of being driven by an intraaortic micro-electric motor, it was powered by a magnetic field from outside of body. To ensure the perfusion of coronary artery, the axial flow pump is to be implanted in the position of aortic valve. It could serve as either a blood pump or a mechanical valve depending on the power input. This research tested the feasibility of the new concept in model study. A column, made from permanent magnet, is jointed to an impeller in a concentric way to form a “rotor-impeller”. Supported by a hanging shaft cantilevered in the center of a rigid cage, the rotor-impeller can be turned by the magnetic field in the surrounding space. In the present prototype, the rotor is 8 mm in diameter and 15 mm in length, the impeller has 3 vanes with an outer diameter of 18 mm. The supporting cage is 22 mm in outer diameter and 20 mm in length. When tested, the DAV prototype is inserted into the tube of a mock circuit. The alternative magnetic field is produced by a rotating magnet placed side by side with the rotor-impeller at a distance of 30 mm. Once the alternative magnetic field is presented in the surrounding space, the DAV starts to turn, leading to a pressure difference and liquid flow in the tube. The flow rate or pressure difference is proportioned to rotary speed. At the maximal output of hydraulic power, the flow rate reached 5 L/min against an afterload of 100 mmHg. The maximal pressure difference generated by DAV at a rotation rate of 12600 r/min was 147 mmHg. The preliminary results demonstrated the feasibility of “DAV”, further research on this concept is justifiable.     

In vitro motility assay of atrial and ventricular myosin from pig

The role of myosin isoforms in determining contractile filament velocity in the atrium and ventricle of the pig heart was studied by measuring the motion of fluorescently labeled actin over myosin (in vitro motility assay). A rapid and relatively simple method for purification of myosin from small tissue samples was used. The relative extent of light chain-2 phosphorylation was about 30% in both atrial and ventricular myosin extracts. Although the extracted myosin was not free from contaminating proteins, mainly actin, the mean velocity at optimal pH and 32°C of both atrial (3.3 μm/s) and ventricular (2.3 μm/s) myosin were similar to those obtained using extensively purified myosin. The filament sliding velocities using isolated myosin and actin are lower than those estimated from previously published experiments on skinned fiber preparations, which might reflect an influence on sliding velocity by the filament organization or regulatory proteins in the muscle fiber. However, the ratio between velocities of atrial and ventricular myosin was similar in the motility assay (1.5) and muscle fiber experiments (1.6), which might suggest that these two methods reflect the same fundamental processes in cardiac contraction and that the difference in filament sliding velocity between the atrium and ventricle of the pig heart is determined my their myosin isoforms. J. Cell. Biochem. 67:241–247, 1997. © 1997 Wiley-Liss, Inc.     

Circulating gonadotropins levels and contribution of different large antral follicles to isofolliculia in sheep

The aim of this study was first to search for isofolliculia in right and left ovaries during postnatal development of Ouled Djellel ewe lambs, a non-seasonal breed of sheep. In addition, the contribution of different sizes of large antral follicles to this phenomenon was studied, and finally the variations in both plasma FSH and LH levels during this period of life were determined. Plasma was collected from groups of four ewe lambs at 0 (<24 h), 1 week, and every 2 weeks from 4 to 14 weeks of age. Thereafter, each group was slaughtered, right and left ovaries recovered, weighed and their length and width measured. One ovary was fixed in Bouin–Holland's solution and prepared for histological study. The other one was immediately frozen and cut in a cryostat and prepared for histochemical study. This latter method was used to detect the activity of alkaline phosphatase, lactate dehydrogenase, NADH₂-tetrasolium reductase, and glucose-6-phosphate dehydrogenase enzymes. Number and size of antral follicles and their contribution to isofolliculia were determined from ovarian sections of both studies. Isofolliculia was seen in right and left ovaries of Ouled Djellel ewe lambs at 4, 6, 8, and 10 weeks of age. This phenomenon was characterized by the presence of large antral follicles almost equal in size and total enzymatic inactivity in the interstitium. Weight and dimensions of right and left ovaries increased rapidly from birth to 4 weeks of age, and then rose gradually to week 8 and then rising again to week 10, followed by a decline at 12 and 14 weeks of age. All large antral follicles contributed to isofolliculia in right and left ovaries but, the percentage of antral follicles <2 mm at 4, 6, 8, and 10 weeks of age were significantly greater than the percentage of follicles ≥2 and <3 mm and the contribution of follicles ≥3 mm was the lowest. FSH levels increased slowly from birth to 6 weeks of age then, increased rapidly to week 10, followed by a decline at weeks 12 and 14. LH was low at birth and the level increased slowly to 8 weeks of age, followed by a further rapid increase at 10 weeks of age. All parameters studied did not show any significant differences between the right and left ovary. It was concluded that isofolliculia occurred between 4 and 10 weeks of age in left and right ovaries of Ouled Djellel ewe lambs. This phenomenon was characterized by the increase of both ovarian weights and dimensions, and of plasma FSH and LH levels. All large antral follicles ≥1 mm in diameter contributed to isofolliculia but the contribution of antral follicles <2 mm was greater than the contribution of antral follicles ≥2.     

Quantification and Proteomic Characterization of β-Hydroxybutyrylation Modification in the Hearts of AMPKα2 Knockout Mice

AMP-activated protein kinase alpha 2 (AMPKα2) regulates energy metabolism, protein synthesis, and glucolipid metabolism myocardial cells. Ketone bodies produced by fatty acid β-oxidation, especially β-hydroxybutyrate, are fatty energy–supplying substances for the heart, brain, and other organs during fasting and long-term exercise. They also regulate metabolic signaling for multiple cellular functions. Lysine β-hydroxybutyrylation (Kbhb) is a β-hydroxybutyrate–mediated protein posttranslational modification. Histone Kbhb has been identified in yeast, mouse, and human cells. However, whether AMPK regulates protein Kbhb is yet unclear. Hence, the present study explored the changes in proteomics and Kbhb modification omics in the hearts of AMPKα2 knockout mice using a comprehensive quantitative proteomic analysis. Based on mass spectrometry (LC-MS/MS) analysis, the number of 1181 Kbhb modified sites in 455 proteins were quantified between AMPKα2 knockout mice and wildtype mice; 244 Kbhb sites in 142 proteins decreased or increased after AMPKα2 knockout (fold change >1.5 or <1/1.5, p < 0.05). The regulation of Kbhb sites in 26 key enzymes of fatty acid degradation and tricarboxylic acid cycle was noted in AMPKα2 knockout mouse cardiomyocytes. These findings, for the first time, identified proteomic features and Kbhb modification of cardiomyocytes after AMPKα2 knockout, suggesting that AMPKα2 regulates energy metabolism by modifying protein Kbhb.