A synteny map of the horse genome comprised of 240 microsatellite and RAPD markers

To generate a domestic horse genome map we integrated synteny information for markers screened on a somatic cell hybrid (SCH) panel with published information for markers physically assigned to chromosomes. The mouse-horse SCH panel was established by fusing pSV2neo transformed primary horse fibroblasts to either RAG or LMTk mouse cells, followed by G418 antibiotic selection. For each of the 108 cell lines of the panel, we defined the presence or absence of 240 genetic markers by PCR, including 58 random amplified polymorphic DNA (RAPD) markers and 182 microsatellites. Thirty-three syntenic groups were defined, comprised of two to 26 markers with correlation coefficient (r) values ranging from 0.70 to 1.0. Based on significant correlation values with physically mapped microsatellite (type II) or gene (type I) markers, 22 syntenic groups were assigned to horse chromosomes (1, 2, 3, 4, 6, 9, 10, 11, 12, 13, 15, 18, 19, 20, 21, 22, 23, 24, 26, 30, X and Y). The other 11 syntenic groups were provisionally assigned to the remaining chromosomes based on information provided by heterologous species painting probes and work in progress with type I markers.     

Presence of leukaemia inhibitory factor (LIF) and LIF-receptor chain (gp190) in osteoclast-like cells cultured from human giant cell tumour of bone. Ultrastructural distribution.

The behaviour of multinucleated giant cells (MNC) obtained from a giant cell tumour of the tibia and cultured on glass coverslips or on devitalized dentin slices was investigated using light and electron microscopy. Cells were studied in the presence or absence of LIF a cytokine known to be involved in bone turnover and to act as a growth factor in some solid tumours. The direct effect of LIF on MNC was examined by a post-embedding colloidal gold immunocytochemistry process using human anti-LIF and anti-LIF-receptor (chain gp190) antibodies. After 7 days of culture, the MNC obtained displayed osteoclast immunocytochemical features. Moreover, these MNC were able to resorb large amounts of dentin and presented typical features of active osteoclasts. Immunolocalization of LIF and LIF-receptor revealed the presence of this cytokine and its receptor within the cytoplasm and nucleus of active resorbing MNC. LIF upmodulated MNC number and nucleation but decreased their ability to resorb dentin. The present study suggests that MNC obtained from human GCT, currently considered as osteoclast-like cells, are targets for LIF and may be a source of LIF production in this pathological condition.     

Effect of resistance training on single muscle fiber contractile function in older men.

The purpose of this study was to examine single cell contractile mechanics of skeletal muscle before and after 12 wk of progressive resistance training (PRT) in older men (n = 7; age = 74 +/- 2 yr and weight = 75 +/- 5 kg). Knee extensor PRT was performed 3 days/wk at 80% of one-repetition maximum. Muscle biopsy samples were obtained from the vastus lateralis before and after PRT (pre- and post-PRT, respectively). For analysis, chemically skinned single muscle fibers were studied at 15 degrees C for peak tension [the maximal isometric force (P(o))], unloaded shortening velocity (V(o)), and force-velocity parameters. In this study, a total of 199 (89 pre- and 110 post-PRT) myosin heavy chain (MHC) I and 99 (55 pre- and 44 post-PRT) MHC IIa fibers were reported. Because of the minimal number of hybrid fibers identified post-PRT, direct comparisons were limited to MHC I and IIa fibers. Muscle fiber diameter increased 20% (83 +/- 1 to 100 +/- 1 microm) and 13% (86 +/- 1 to 97 +/- 2 microm) in MHC I and IIa fibers, respectively (P < 0.05). P(o) was higher (P < 0.05) in MHC I (0.58 +/- 0.02 to 0.90 +/- 0.02 mN) and IIa (0.68 +/- 0.02 to 0.85 +/- 0.03 mN) fibers. Muscle fiber V(o) was elevated 75% (MHC I) and 45% (MHC IIa) after PRT (P < 0.05). MHC I and IIa fiber power increased (P < 0.05) from 7.7 +/- 0.5 to 17.6 +/- 0.9 microN. fiber lengths. s(-1) and from 25.5 to 41.1 microN. fiber lengths. s(-1), respectively. These data indicate that PRT in elderly men increases muscle cell size, strength, contractile velocity, and power in both slow- and fast-twitch muscle fibers. However, it appears that these changes are more pronounced in the MHC I muscle fibers.     

Synthesis and biological evaluation of new donepezil-like Thiaindanones as AChE inhibitors

Pharmacomodulations of previously reported thiaindanones related to donepezil were achieved with the aim to enhance their AChE inhibitory activity. Condensation of the cyclopentane carbonyl group into hydrazone or cyanolefine derivatives, as well as its hydrogenation and the subsequent substitution of the resulting hydroxyl group led to new 2-(4-benzylpiperazin-1-yl)-N-(1,3-dibromo-6-hydroxy-5,6-dihydro-4H-cyclopenta[c]thien-4-yl) acetamides. The in vitro evaluation of this new series, according to the method of Ellman, shows however that it conserved only partially the biological activity. The best compound remains the alcohol 11 (IC(50) = 0.40 microM, against 0.02 microM for donepezil).     

Assembly-dependent translation of subunits 6 (Atp6) and 9 (Atp9) of ATP synthase in yeast mitochondria

The yeast mitochondrial ATP synthase is an assembly of 28 subunits of 17 types of which 3 (subunits 6, 8, and 9) are encoded by mitochondrial genes, while the 14 others have a nuclear genetic origin. Within the membrane domain (F_O) of this enzyme, the subunit 6 and a ring of 10 identical subunits 9 transport protons across the mitochondrial inner membrane coupled to ATP synthesis in the extra-membrane structure (F₁) of ATP synthase. As a result of their dual genetic origin, the ATP synthase subunits are synthesized in the cytosol and inside the mitochondrion. How they are produced in the proper stoichiometry from two different cellular compartments is still poorly understood. The experiments herein reported show that the rate of translation of the subunits 9 and 6 is enhanced in strains with mutations leading to specific defects in the assembly of these proteins. These translation modifications involve assembly intermediates interacting with subunits 6 and 9 within the final enzyme and cis-regulatory sequences that control gene expression in the organelle. In addition to enabling a balanced output of the ATP synthase subunits, these assembly-dependent feedback loops are presumably important to limit the accumulation of harmful assembly intermediates that have the potential to dissipate the mitochondrial membrane electrical potential and the main source of chemical energy of the cell.     

High and low affinity receptors for human interleukin for DA cells/leukemia inhibitory factor on human cells. Molecular characterization and cellular distribution.

Radioiodinated recombinant human interleukin DA (HILDA)/leukemia inhibitory factor (LIF) purified from conditioned medium of Chinese hamster ovary transfected cells enabled the identification of specific receptor sites on a variety of human cell types. Using low concentrations (up to 500 pM) of the ligand iodinated at a high specific radioactivity, high affinity receptors (equilibrium dissociation constant Kd in the range of 30-100 pM) were first demonstrated. They were expressed at low levels by human peripheral blood monocytes but not by lymphocytes, NK cells, granulocytes, and platelets. The myelomonocytic cell line THP1 as well as the T lymphoma cell line HSB2 and the lymphoblastoid B cell line DAB were also receptor-negative. In contrast, most of the non-lymphoid tumoral cell lines tested, including melanomas, neuroblastomas, and carcinomas, expressed high affinity HILDA/LIF receptors at variable levels (Bmax from 20 to 600 sites/cell). The kinetics of HILDA/LIF high affinity binding to the choriocarcinoma JAR cell line were characterized at 4 degrees C with association and dissociation rate constants of k1 = 2.2 10(9) M-1 min-1 and k-1 = 0.0084 min-1, respectively, corresponding to a steady-state dissociation constant k1/k-1 = 3.8 pM. The subsequent use of higher concentrations of HILDA/LIF labeled at a lower specific radioactivity enabled the identification of a low affinity component on several cell lines (Kd in the range of 1-4 nM; Bmax from 1,000 to 5,000 sites/cell). On JAR cells, this low affinity component was characterized by association and dissociation rate constants at 4 degrees C of k1 = 7.3 10(7) M-1 min-1 and k-1 = 0.19 min-1, respectively (k-1/k1 = 2.6 nM). Affinity cross-linking of HILDA/LIF to JAR cells showed two cross-linked species under both reducing and nonreducing conditions corresponding to receptor species of 120 and 250 kDa, respectively. Whereas both bands had similar intensities under high affinity conditions, the higher band predominated under low affinity conditions. Our data suggest that the 250-kDa chain could constitute the low affinity binding component whereas the association of both 250- and 120-Da subunits would form the high affinity structure.