Effects of combined centrifugation and ultraviolet irradiation of eggs on pattern formation in Chironomus embryos

Summary

Combined mild centrifugation and uv irradiation of Chironomus embryos modified the developmental types expected from centrifugation alone, somewhat differently from the combined strong centrifugation and uv irradiation of Smittia embryos. The modifications changed with the stages irradiated. The change caused by anterior irradiation may depend on whether or not a part of the cytoplasmic zone is irradiated simultaneously with the anterior yolky end; because most of the cytoplasm lies in the posterior half of egg at early irradiation, while the tip of the cytoplasm redistributes near the anterior end by the late irradiation. Early uv irradiation of the anterior end of centrifuged eggs, causing the formation of a double abdomen (DA) or an inverted embryo, is not photoreversible, while the uv damage to the anterior end of uncentrifuged eggs, inducing DA, is. These facts suggest that there is another photoirreversible uv target in addition to the photoreversible target for DA induction or the anterior determinant shown in Smittia. Other changes, such as the induction of a double cephalon by late irradiation of the centrifuged egg, are photoreversible, but in an unusual way in that the level of photorecovery is similar to the result of incubation in the dark after early irradiation, and not to that of the centrifuged controls. These modified results were then compared with those for Smittia embryos.     

Circadian Phase-Dependent Protein and Tissular Binding of Three Local Anesthetics (Bupivacaine,Etidocaine, and Mepivacaine) in Mice: A Possible Mechanismof Their Chronotoxicokinetics?

The aim of this study was to investigate the possible influence of the time of administration on bupivacaine (B), etidocaine (E), and mepivacaine (M) protein and tissue (brain and heart) binding. For each anesthetic agent, a single dose of B (20 mg/kg), E (40 mg/kg), or M (60 mg/kg) was administered intraperito-neally at 10:00,16:00,22:00, and 04:00 h. Blood and tissue samples were collected 15 min after drug administration. This study documents significant circadian variations in protein and tissue binding of the three local anesthetic agents. We did not demonstrate a temporal relationship between the respective free and tissue levels. Thus, the temporal variations of free plasma, brain, and heart levels do not seem to be involved in the temporal changes of induced mortality.     

First record of dicephalia in a bull shark Carcharhinus leucas (Chondrichthyes: Carcharhinidae) foetus from the Gulf of Mexico,U.S.A.

The first recorded incidence of dicephalia in a bull shark Carcharhinus leucas is reported from a foetus collected by a fisherman in the Gulf of Mexico near Florida, U.S.A. External examination, Radiography and magnetic resonance imaging revealed a case of monosomic dicephalia where the axial skeleton and internal organs were found to divide into parallel systems anterior to the pectoral girdle resulting in two well‐developed heads.     

Cardiomyopathy Mutations in the Tail of β-Cardiac Myosin Modify the Coiled-coil Structure and Affect Integration into Thick Filaments in Muscle Sarcomeres in Adult Cardiomyocytes

It is unclear why mutations in the filament-forming tail of myosin heavy chain (MHC) cause hypertrophic or dilated cardiomyopathy as these mutations should not directly affect contraction. To investigate this, we first investigated the impact of five hypertrophic cardiomyopathy-causing (N1327K, E1356K, R1382W, E1555K, and R1768K) and one dilated cardiomyopathy-causing (R1500W) tail mutations on their ability to incorporate into muscle sarcomeres in vivo. We used adenoviral delivery to express full-length wild type or mutant enhanced GFP-MHC in isolated adult cardiomyocytes. Three mutations (N1327K, E1356K, and E1555K) reduced enhanced GFP-MHC incorporation into muscle sarcomeres, whereas the remainder had no effect. No mutations significantly affected contraction. Fluorescence recovery after photobleaching showed that fluorescence recovery for the mutation that incorporated least well (N1327K) was significantly faster than that of WT with half-times of 25.1 ± 1.8 and 32.2 ± 2.5 min (mean ± S.E.), respectively. Next, we determined the effects of each mutation on the helical properties of wild type and seven mutant peptides (7, 11, or 15 heptads long) from the myosin tail by circular dichroism. R1382W and E1768K slightly increased the α-helical nature of peptides. The remaining mutations reduced α-helical content, with N1327K showing the greatest reduction. Only peptides containing residues 1301–1329 were highly α-helical suggesting that this region helps in initiation of coiled coil. These results suggest that small effects of mutations on helicity translate into a reduced ability to incorporate into sarcomeres, which may elicit compensatory hypertrophy.     

A Structural View on the Functional Importance of the Sugar Moiety and Steroid Hydroxyls of Cardiotonic Steroids in Binding to Na,K-ATPase

The Na,K-ATPase is specifically inhibited by cardiotonic steroids (CTSs) like digoxin and is of significant therapeutic value in the treatment of congestive heart failure and arrhythmia. Recently, new interest has arisen in developing Na,K-ATPase inhibitors as anticancer agents. In the present study, we compare the potency and rate of inhibition as well as the reactivation of enzyme activity following inhibition by various cardiac glycosides and their aglycones at different pH values using shark Na,K-ATPase stabilized in the E2MgP_i or in the E2BeF_x conformations. The effects of the number and nature of various sugar residues as well as changes in the positions of hydroxyl groups on the β-side of the steroid core of cardiotonic steroids were investigated by comparing various cardiac glycoside compounds like ouabain, digoxin, digitoxin, and gitoxin with their aglycones. The results confirm our previous hypothesis that CTS binds primarily to the E2-P ground state through an extracellular access channel and that binding of extracellular Na⁺ ions to K⁺ binding sites relieved the CTS inhibition. This reactivation depended on the presence or absence of the sugar moiety on the CTS, and a single sugar is enough to impede reactivation. Finally, increasing the number of hydroxyl groups of the steroid was sterically unfavorable and was found to decrease the inhibitory potency and to confer high pH sensitivity, depending on their position on the steroid β-face. The results are discussed with reference to the recent crystal structures of Na,K-ATPase in the unbound and ouabain-bound states.     

Tbx18 regulates development of the epicardium and coronary vessels

The epicardium and coronary vessels originate from progenitor cells in the proepicardium. Here we show that Tbx18, a T-box family member highly expressed in the proepicardium, controls critical early steps in coronary development. In Tbx18^−/− mouse embryos, both the epicardium and coronary vessels exhibit structural and functional defects. At E12.5, the Tbx18-deficient epicardium contains protrusions and cyst-like structures overlying a disorganized coronary vascular plexus that contains ectopic structures resembling blood islands. At E13.5, the left and right coronary stems form correctly in mutant hearts. However, analysis of PECAM-1 whole mount immunostaining, distribution of SM22α^lacZ/+ activity, and analysis of coronary vascular casts suggest that defective vascular plexus remodeling produces a compromised arterial network at birth consisting of fewer distributing conduit arteries with smaller lumens and a reduced capacity to conduct blood flow. Gene expression profiles of Tbx18^−/− hearts at E12.5 reveal altered expression of 79 genes that are associated with development of the vascular system including sonic hedgehog signaling components patched and smoothened, VEGF-A, angiopoietin-1, endoglin, and Wnt factors compared to wild type hearts. Thus, formation of coronary vasculature is responsive to Tbx18-dependent gene targets in the epicardium, and a poorly structured network of coronary conduit vessels is formed in Tbx18 null hearts due to defects in epicardial cell signaling and fate during heart development. Lastly, we demonstrate that Tbx18 possesses a SRF/CArG box dependent repressor activity capable of inhibiting progenitor cell differentiation into smooth muscle cells, suggesting a potential function of Tbx18 in maintaining the progenitor status of epicardial-derived cells.     

8-Oxoguanine DNA glycosylase 1 (ogg1) maintains the function of cardiac progenitor cells during heart formation in zebrafish

Genomic damage may devastate the potential of progenitor cells and consequently impair early organogenesis. We found that ogg1, a key enzyme initiating the base-excision repair, was enriched in the embryonic heart in zebrafish. So far, little is known about DNA repair in cardiogenesis. Here, we addressed the critical role of ogg1 in cardiogenesis for the first time. ogg1 mainly expressed in the anterior lateral plate mesoderm (ALPM), the primary heart tube, and subsequently the embryonic myocardium by in situ hybridisation. Loss of ogg1 resulted in severe cardiac morphogenesis and functional abnormalities, including the short heart length, arrhythmia, decreased cardiomyocytes and nkx2.5⁺ cardiac progenitor cells. Moreover, the increased apoptosis and repressed proliferation of progenitor cells caused by ogg1 deficiency might contribute to the heart phenotype. The microarray analysis showed that the expression of genes involved in embryonic heart tube morphogenesis and heart structure were significantly changed due to the lack of ogg1. Among those, foxh1 is an important partner of ogg1 in the cardiac development in response to DNA damage. Our work demonstrates the requirement of ogg1 in cardiac progenitors and heart development in zebrafish. These findings may be helpful for understanding the aetiology of congenital cardiac deficits.     

Balloon Venoplasty of Subclavian Vein and Brachiocephalic Junction to Enable Left Ventricular Lead Placement for Cardiac Resynchronisation Therapy

This report describes the successful implantation of a LV lead using balloon venoplasty to overcome a very tight stenosis of the right subclavian vein / brachiocephalic junction for cardiac resynchronisation therapy (CRT-P) in a patient with a right sided CRT-P system and a failed epicardial LV lead. It is important for device implanters to be familiar with interventional equipments and techniques such as balloon venoplasty to overcome difficult venous access.