Assessment of Genetic Variability of Fish Personality Traits using Rainbow Trout Isogenic Lines

The study of inter-individual variability of personality in fish is a growing field of interest but the genetic basis of this complex trait is still poorly investigated due to the difficulty in controlling fish genetic origin and life history. When available, isogenic lines that allow performing independent tests on different individuals having identical genotype constitute a very relevant experimental material to disentangle the genetic and environmental components of behavioural individuality. We took advantage of heterozygous isogenic lines to investigate the personality in rainbow trout through the analysis of their reactions to different experimental situations. To this end, seven to ten rainbow trout isogenic lines were screened for their spatial exploratory behaviour, their flight response toward a stressor and their risk taking behaviour. Results showed that some lines seemed less sensitive to new events or environmental changes and could be defined as low responsive, while others were very sensitive and defined as high responsive. The use of isogenic lines highlighted the importance of genetic factors, in combination with life history, in the expression of personality in domesticated fish.     

CLCNKB mutations causing mild Bartter syndrome profoundly alter the pH and Ca2+ dependence of ClC-Kb channels

ClC-Kb, a member of the ClC family of Cl^? channels/transporters, plays a major role in the absorption of NaCl in the distal nephron. CLCNKB mutations cause Bartter syndrome type 3, a hereditary renal salt-wasting tubulopathy. Here, we investigate the functional consequences of a Val to Met substitution at position 170 (V170M, α helix F), which was detected in eight patients displaying a mild phenotype. Conductance and surface expression were reduced by ~40–50 %. The regulation of channel activity by external H⁺ and Ca²⁺ is a characteristic property of ClC-Kb. Inhibition by external H⁺ was dramatically altered, with pK_H shifting from 7.6 to 6.0. Stimulation by external Ca²⁺ on the other hand was no longer detectable at pH 7.4, but was still present at acidic pH values. Functionally, these regulatory modifications partly counterbalance the reduced surface expression by rendering V170M hyperactive. Pathogenic Met170 seems to interact with another methionine on α helix H (Met227) since diverse mutations at this site partly removed pH sensitivity alterations of V170M ClC-Kb. Exploring other disease-associated mutations, we found that a Pro to Leu substitution at position 124 (α helix D, Simon et al., Nat Genet 1997, 17:171–178) had functional consequences similar to those of V170M. In conclusion, we report here for the first time that ClC-Kb disease-causing mutations located around the selectivity filter can result in both reduced surface expression and hyperactivity in heterologous expression systems. This interplay must be considered when analyzing the mild phenotype of patients with type 3 Bartter syndrome.     

Spermatogonia differentiation requires retinoic acid receptor γ

Vitamin A is instrumental to mammalian reproduction. Its metabolite, retinoic acid (RA), acts in a hormone-like manner through binding to and activating three nuclear receptor isotypes, RA receptor (RAR)α (RARA), RARβ, and RARγ (RARG). Here, we show that 1) RARG is expressed by A aligned (A(al)) spermatogonia, as well as during the transition from A(al) to A(1) spermatogonia, which is known to require RA; and 2) ablation of Rarg, either in the whole mouse or specifically in spermatogonia, does not affect meiosis and spermiogenesis but impairs the A(al) to A(1) transition in the course of some of the seminiferous epithelium cycles. Upon ageing, this phenomenon yields seminiferous tubules containing only spermatogonia and Sertoli cells. Altogether, our findings indicate that RARG cell-autonomously transduces, in undifferentiated spermatogonia of adult testes, a RA signal critical for spermatogenesis. During the prepubertal spermatogenic wave, the loss of RARG function can however be compensated by RARA, as indicated by the normal timing of appearance of meiotic cells in Rarg-null testes. Accordingly, RARG- and RARA-selective agonists are both able to stimulate Stra8 expression in wild-type prepubertal testes. Interestingly, inactivation of Rarg does not impair expression of the spermatogonia differentiation markers Kit and Stra8, contrary to vitamin A deficiency. This latter observation supports the notion that the RA-signaling pathway previously shown to operate in Sertoli cells also participates in spermatogonia differentiation.     

Mineralocorticoid receptor overexpression facilitates differentiation and promotes survival of embryonic stem cell-derived neurons

Mineralocorticoid receptor (MR), highly expressed in the hippocampus, binds corticosteroid hormones and coordinately participates, with the glucocorticoid receptor, to the control of stress responses, memorization, and behavior. To investigate the impact of MR in neuronal survival, we generated murine embryonic stem (ES) cells that overexpress human MR (hMR) (P1-hMR) and are induced to differentiate into mature neurons. We showed that recombinant MR expression increased throughout differentiation and is 2-fold higher in P1-hMR ES-derived neurons compared with wild-type controls, whereas glucocorticoid receptor expression was unaffected. Although proliferation and early neuronal differentiation were comparable in P1-hMR and wild-type ES cells, MR overexpression was associated with higher late neuronal marker expression (microtubule-associated protein 2 and β-tubulin III). This was accompanied by a shift towards neuron survival with an increased ratio of anti- vs. proapoptotic molecules and 50% decreased caspase 3 activity. Knocking down MR overexpression by small interfering RNA drastically reversed neuroprotective effects with reduced Bcl(2)/Bax ratio and decreased microtubule-associated protein 2 expression. P1-hMR neurons were protected against oxidative stress-induced apoptosis through reduced caspase 3 activation and drastically increased Bcl(2)/Bax ratio and β-tubulin III expression. We demonstrated the involvement of MR in neuronal differentiation and survival and identify MR as an important neuroprotective mediator opening potential pharmacological strategies.     

Retinoic acid induces Sertoli cell paracrine signals for spermatogonia differentiation but cell autonomously drives spermatocyte meiosis

Direct evidence for a role of endogenous retinoic acid (RA), the active metabolite of vitamin A in the initial differentiation and meiotic entry of spermatogonia, and thus in the initiation of spermatogenesis is still lacking. RA is synthesized by dedicated enzymes, the retinaldehyde dehydrogenases (RALDH), and binds to and activates nuclear RA receptors (RARA, RARB, and RARG) either within the RA-synthesizing cells or in the neighboring cells. In the present study, we have used a combination of somatic genetic ablations and pharmacological approaches in vivo to show that during the first, prepubertal, spermatogenic cycle (i) RALDH-dependent synthesis of RA by Sertoli cells (SC), the supporting cells of the germ cell (GC) lineage, is indispensable to initiate differentiation of A aligned into A1 spermatogonia; (ii) RARA in SC mediates the effects of RA, possibly through activating Mafb expression, a gene whose Drosophila homolog is mandatory to GC differentiation; (iii) RA synthesized by premeiotic spermatocytes cell autonomously induces meiotic initiation through controlling the RAR-dependent expression of Stra8. Furthermore, we show that RA of SC origin is no longer necessary for the subsequent spermatogenic cycles but essential to spermiation. Altogether, our data establish that the effects of RA in vivo on spermatogonia differentiation are indirect, via SC, but direct on meiotic initiation in spermatocytes, supporting thereby the notion that, contrary to the situation in the female, RA is necessary to induce meiosis in the male.     

Early matched sibling hematopoietic cell transplantation for adult AML in first remission using an age-adapted strategy: long-term results of a prospective GOELAMS study

The LAM2001 phase 3 trial, involving 832 patients with acute myeloid leukemia (AML; median: 46 years) proposed HLA-identical sibling allograft HSCT for all patients with an identified donor. The trial compared reduced-intensity conditioning (RIC) for patients older than 50 years of age (N = 47) and myeloablative conditioning for younger patients (N = 117). BM HSCT was performed in the younger patients, while the older ones received a consolidation course, followed by peripheral blood allo-HSCT using RIC. The incidence of grade II-IV acute GVHD, was 51.9% (95% confidence interval [CI]: 42.1-61.8) and 11.3% (1.6-21.2) after myeloablative or RIC, respectively (P < .0001) and that of chronic GVHD 45.8% (95% CI: 34.8-56.7) and 41.7% (24.7-58.6; NS). Cumulative incidence of nonrelapse mortality at 108 months was 15.8% (95% CI: 9.8-23.2) for myeloablative, and 6.5% (0.2-16.2) for RIC (NS). CI of relapse at 108 months was 21.7% (95% CI: 13.9-28.6) and 28.6% (16.5-43.4; NS). Overall survival at 108 months was 63.4% (95% CI: 54.6-72.2) and 65.8% (52.2-72.2), respectively, after myeloablative or RIC (NS). RIC peripheral blood stem cell allo-HSCT is prospectively feasible for patients between the ages of 51 and 60 years without excess of relapse or nonrelapse mortality, and compares favorably with myeloablative marrow allo-HSCT proposed to younger patients.