Purging of acute myeloid leukemic cells by ether lipids and hyperthermia

Ether lipids (EL) and hyperthermia have been shown to possess a relatively selective cytotoxicity to leukemic cells. In this study, the combined effects of EL (ET-18-OCH3, ET-16-NHCOCH3, or BM 41.440) and hyperthermia on the growth of hematopoietic progenitors, myeloid leukemic cell lines, and leukemic cells obtained from patients with acute myeloid leukemia (AML) were examined to determine if this combination resulted in a greater selective killing of leukemic cells than that achieved by either EL or heat alone. When the cells were treated simultaneously with EL (50 micrograms/mL) and hyperthermia (42 degrees C) for one hour, the killing of leukemic cell line cells was enhanced considerably. Among the three EL, however, the combination of ET-18-OCH3 and heat seemed to be the most cytotoxic to leukemic cell line cells with no effect on the growth of hematopoietic progenitors. An increase in the duration of treatment with ET-18-OCH3 to four hours with heat added during the last hour resulted in a further reduction of leukemic cell line cells while sparing 50% of hematopoietic progenitors after cryopreservation. The combined treatment with ET-18-OCH3 and heat also inhibited the growth of leukemic progenitors obtained from AML patients by 97% to 100%. These data indicate that the combined treatment with EL and hyperthermia might offer an efficient means to eliminate myeloid leukemic cells in vitro.     

National clinical guidelines for management of the palatally ectopic maxillary canine

This review summarises updated clinical guidelines produced by the Clinical Standards Committee of the Faculty of Dental Surgery, Royal College of Surgeons of England (FDSRCS). This guideline on the management of the palatally ectopic maxillary canine illustrates the information contained in the recently updated online version. The timely recognition of ectopic canines is important for the overall management of the dentition. This review illustrates five management strategies for ectopic permanent canines: interceptive treatment by extraction of the deciduous canine, surgical exposure and orthodontic alignment, surgical removal of the palatally ectopic permanent canine, auto-transplantation and no active treatment/leave and observe. The current available evidence for each of these management options has been evaluated and awarded a grade used by the Scottish Intercollegiate Guidelines Network.     

GABAA receptors can initiate the formation of functional inhibitory GABAergic synapses

The mechanisms that underlie the selection of an inhibitory GABAergic axon's postsynaptic targets and the formation of the first contacts are currently unknown. To determine whether expression of GABA_A receptors (GABA_ARs) themselves – the essential functional postsynaptic components of GABAergic synapses – can be sufficient to initiate formation of synaptic contacts, a novel co‐culture system was devised. In this system, the presynaptic GABAergic axons originated from embryonic rat basal ganglia medium spiny neurones, whereas their most prevalent postsynaptic targets, i.e. α1/β2/γ2‐GABA_ARs, were expressed constitutively in a stably transfected human embryonic kidney 293 (HEK293) cell line. The first synapse‐like contacts in these co‐cultures were detected by colocalization of presynaptic and postsynaptic markers within 2 h. The number of contacts reached a plateau at 24 h. These contacts were stable, as assessed by live cell imaging; they were active, as determined by uptake of a fluorescently labelled synaptotagmin vesicle‐luminal domain‐specific antibody; and they supported spontaneous and action potential‐driven postsynaptic GABAergic currents. Ultrastructural analysis confirmed the presence of characteristics typical of active synapses. Synapse formation was not observed with control or N‐methyl‐d ‐aspartate receptor‐expressing HEK293 cells. A prominent increase in synapse formation and strength was observed when neuroligin‐2 was co‐expressed with GABA_ARs, suggesting a cooperative relationship between these proteins. Thus, in addition to fulfilling an essential functional role, postsynaptic GABA_ARs can promote the adhesion of inhibitory axons and the development of functional synapses.     

An inhibitory monoclonal antibody to factor X that blocks prothrombin activation but not prothrombinase enzyme assembly

A monoclonal antibody (designated alpha BFX-2b) prepared against bovine factor X inhibited factor X activity in human, bovine, porcine, rabbit, and canine plasma. In assays using purified prothrombinase components, factor Xa, factor Va, phospholipid vesicles, and calcium ion with the fluorescent active site thrombin inhibitor dansylarginyl-N-(3-ethyl-1,5- pentanediyl)amide, the antibody inhibited the conversion of prothrombin to thrombin. Antibody alpha BFX-2b also blocked prothrombinase cleavage of the macromolecular substrates prethrombin 1 and prethrombin 2 but did not inhibit factor Xa hydrolysis of the synthetic substrate benzoyl- Ile-Glu-Gly-Arg-p-nitroanilide. The antibody also prevented the inactivation of factor Xa by antithrombin III but did not prevent the inactivation by soybean trypsin inhibitor. Antibody alpha BFX-2b bound factor Xa with a stoichiometry of 1:1 and an apparent dissociation constant of 9.0 x 10(-11) mol/L as estimated from its inhibition of prothrombinase activity. Antibody alpha BFX-2b did not prevent binding of factor Xa to factor Va-phospholipid as measured by using fluorescence polarization or high-pressure liquid gel chromatography with the fluorescent Factor Xa analogue dansyl-glutamyl-glycyl-arginyl- Xa. Immunoblotting of factor X following electrophoresis on sodium dodecyl sulphate-polyacrylamide gels and transfer to nitrocellulose indicated that the antigenic determinant recognized by antibody alpha BFX-2b was found on the heavy chain of factors X and Xa. From these observations it can be concluded that antibody alpha BFX-2b recognizes a highly conserved epitope on the factor X heavy chain that is remote from the topographic sites required for prothrombinase complex assembly and substrate hydrolysis but may be located at or near a portion of the macromolecular substrate binding site.     

Platelet-collagen interaction: inhibition by ristocetin and enhancement by von Willebrand factor-platelet binding

The contribution of von Willebrand factor (vWF)-platelet binding to platelet-collagen interaction was examined in vitro. The binding of vWF to platelets was mediated and regulated by ristocetin. Subthreshold concentrations of ristocetin (less than or equal to 1 mg/mL), insufficient to cause ristocetin-induced platelet aggregation (RIPA), were added to platelet-rich plasma (PRP) prior to the addition of collagen. The collagen-induced platelet aggregation (CIPA) was modified by ristocetin and the degree of alteration was dependent on the ristocetin concentration. Response as a function of ristocetin concentration was designated the Collagen-Platelet Aggregation Response (CoI-PAR). In normal PRP the CoI-PAR was a progressive inhibition followed by decreasing inhibition and then an enhanced response. The enhanced response occurred over a narrow range of ristocetin concentrations (0.8 to 1.0 mg/mL). In the absence of vWF (severe von Willebrand's disease, Type I, vWF less than 1%) the CoI-PAR was a progressive, eventually complete inhibition with no enhanced response (with ristocetin concentrations up to 3.0 mg/mL). With addition of vWF to this PRP an enhanced response was observed at a ristocetin concentration inversely proportional to the vWF level. PRP from a patient with severe Hemophilia A showed a response within the normal range. Subthreshold ristocetin did not cause plasma protein precipitation or platelet release of 3H-serotonin, nor induce micro platelet aggregate formation. Digestion of platelet membrane glycoproteins (GP(s] with chymotrypsin demonstrated that upon removal of GPI, RIPA was absent, CIPA retained and the CoI-PAR was progressive inhibition, with no enhancement. With removal of GPs I, II, and III, RIPA, CIPA, and the CoI-PAR were absent. A dose-response 125I-vWF- platelet binding occurred with increasing ristocetin concentrations which was unchanged by the addition of collagen. These results demonstrated that ristocetin-platelet association inhibited CIPA, and vWF-platelet binding enhanced platelet-collagen adhesion and platelet aggregation. The in vitro-enhanced CIPA represents a vWF-dependent aggregation of sufficient magnitude to overcome the inhibitory effect of ristocetin. These studies demonstrate an influential interaction of ristocetin, vWF, and collagen with the platelet membrane and imply an important hemostatic contribution of vWF-platelet binding in platelet- collagen interaction.     

Acetylcholine receptors at neuromuscular synapses: phylogenetic differences detected by snake alpha-neurotoxins.

Phylogenetic differences in acetylcholine receptors from skeletal neuromuscular synapses of various species of snakes and lizards have been investigated, using the snake venom alpha-neurotoxins alpha-atratoxin (cobrotoxin) and alpha-bungarotoxin. The acetylcholine receptors of the phylogenetically primitive lizards, like those from all other vertebrates previously tested, are blocked by these alpha-neurotoxins. In contrast, receptors from snakes and advanced lizards are insensitive to one or both of the toxins. It is suggested that toxin-resistant acetylcholine receptors appeared early in the evolution of Squamata and preceded the appearance of alpha-neurotoxins.     

Muscular adaptations and insulin‐like growth factor‐1 responses to resistance training are stretch‐mediated

Introduction: Modulation of muscle characteristics was attempted through altering muscle stretch during resistance training. We hypothesized that stretch would enhance muscle responses. Methods: Participants trained for 8 weeks, loading the quadriceps in a shortened (SL, 0–50° knee flexion; n = 10) or lengthened (LL, 40–90°; n = 11) position, followed by 4 weeks of detraining. Controls (CON; n = 10) were untrained. Quadriceps strength, vastus lateralis architecture, anatomical cross‐sectional area (aCSA), and serum insulin‐like growth factor‐1 (IGF‐1) were measured at weeks 0, 8, 10, and 12. Results: Increases in fascicle length (29 ± 4% vs. 14 ± 4%), distal aCSA (53 ± 12% vs. 18 ± 8%), strength (26 ± 6% vs. 7 ± 3%), and IGF‐1 (31 ± 6% vs. 7 ± 6%) were greater in LL compared with SL muscles (P < 0.05). No changes occurred in CON. Detraining decrements in strength and aCSA were greater in SL than LL muscles (P < 0.05). Conclusions: Enhanced muscle in vivo (and somewhat IGF‐1) adaptations to resistance training are concurrent with muscle stretch, which warrants its inclusion within training. Muscle Nerve 49 : 108–119, 2014