Identification of limb-specific Lmx1b auto-regulatory modules with Nail-Patella Syndrome pathogenicity

Resumen del trabajo presentado en el 17th Spanish Society for Developmental Biology Meeting Virtual Meeting, celebrado en modalidad virtual del 18 al 20 de noviembre de 2020.Lmx1b is a LIM homeodomain transcription factor that plays essential roles in a wide range of developmental processes including the development of the kidney, the eye, the dopaminergic and serotonergic neurons and the limb. In the limb, Lmx1b expression is restricted to the dorsal mesenchyme and is responsible for limb dorsalization. Mice lacking functional Lmx1b die at birth because of multisystemic malformations. Homozygous Lmx1b mutants develop a double-ventral limb phenotype with loss of dorsal structures such as the patella, whereas the ventral ones such as the sesamoid bones are duplicated and dorsal tendons and muscles are transformed into mirror images of ventral ones. At superficial level, hair and nails are absent while typical ventral features such as pads are also duplicated. Herein, we report on two conserved Lmx1bassociated cis-regulatory modules (LARM1 and LARM2) located upstream of Lmx1b that are bound by Lmx1b and mediate an autoregulatory loop required to provide adequate levels of Lmx1b expression necessary for limb dorsalization. Combined removal of both enhancer lead to a limb restricted phenotype identical to that of the Lmx1b Knockout mice with no other Lmx1b-related systemic defects, indicating that they are limb specific enhancers. Unexpectedly, the CRISPR/Cas9 deletion of each individual enhancer produced a double ventral phenotype clearly restricted to the posterior or anterior half of the autopod in LARM1 and LARM2 respectively, unveiling spatial modularity in the transcriptional control of Lmx1b in the limb. In humans, LMX1B haploinsufficiency causes Nail-patella syndrome (NPS; MIM 161200), characterized by nail dysplasia, absent/hypoplastic patellae, chronic kidney disease, and glaucoma. Of most interest, we report on two NPS patient families with normal LMX1B coding sequence and limb-restricted phenotype, but with loss-of-function variations in the LARM1/2 region, stressing the role of regulatory modules in disease pathogenesis

Functional analysis of young patients with desmoid-type fibromatosis: Initial surveillance does not jeopardize long term quality of life

International audienceBackground: With recent conservative strategies, prognosis of patients with desmoid-type fibromatosis (DTF) is about function preservation. We analyzed the long-term quality of life (QoL) of pediatric patients with DTF.Methods: All French young patients (<21years) treated between 2005 and 2016 for a DTF in the EpSSG NRSTS-05 study were analyzed. A first wait-and-see strategy was recommended. Patients' QoL was analyzed with the internationally validated Child Health Questionnaire (CHQ). We focused on the relevant subscales scores: physical functioning (PF), role social limitations physical (RP), bodily pain (BP), general health perception (GH) and physical (PhS) and psychosocial (PsS) summary measures.Results: Among the 81 patients, 52 families answered the CHQ (median delay since diagnosis = 6.2years; min2.2-max13.3 years). Median age at diagnosis was 11.5 years. Primary site: limbs (52%), head/neck (27%), or trunk (21%). Five year-Progression Free Survival was 39.1% (95%CI: 27.7-50.5%). As initial management for these 52 patients, 30 patients were first observed (57%), 13 had surgery (25%) and 9 received chemotherapy (18%). Total burden of therapy was exclusive surgery (9pts/18%), exclusive chemotherapy (18pts/35%), surgery + chemotherapy (13pts/25%), chemotherapy + radiotherapy (1 pt), surgery + chemotherapy + radiotherapy (1 pt), wait and see (10 pt). Regarding the parent forms, patients have significant lower PF (86.0vs.96.1; p = 0.03), RP (82.0vs.93.6; p = 0.04), GH (60vs.73; p < 0.005) and PhS (46.2 vs.53; p = 0.02) scores compared to healthy population. Comparison of QoL subscales scores according to initial strategy (wait-and-see vs.surgery/chemotherapy) did not reveal any difference (PF = 87.3vs.84.9; p = 0.80/RP = 83.4vs.78.7; p = 0.72/BP = 78.9vs.78.2; p = 0.95/GH = 59.7vs60; p = 0.97). Similar results were found using the children or adult forms.Conclusions: Initial wait-and-see strategy does not affect long term functional impairment

Normal bone growth requires optimal estrogen levels: negative effects of both high and low dose estrogen on the number of growth plate chondrocytes

Endochondral bone formation at epiphyseal growth plate consists of the synchronized processes of chondrogenesis and cartilage ossification. Estrogen, the major female sex hormone, plays an important role in this process, particularly during the pubertal growth spurt. However, its effects on the growth plate are not completely understood. The aims of this study were to clarify the effects of estrogen on the kinetics of chondrocytes in the growth plates of 10- to 25-week-old female rabbits by studying the effects of ovariectomy or high-dose administration of estrogen on the balance between cell proliferation and death. Forty-eight Japanese white rabbits were divided into three groups: sham operated, ovariectomized, or ovariectomized with subsequent weekly injection of high dose estrogen from 10 weeks. The chondrocyte kinetics was investigated by histomorphometry and immunohistochemistry, using antibodies for caspase-3, a marker of apoptosis, and for proliferating cell nuclear antigen. Both ovariectomized and estrogen-injected rabbits showed a declination of the chondrocyte number although the latter animals indicated a more dramatic effect. Estrogen-injected rabbits showed a decrease in the cell proliferating ability together with an increase in chondrocytes undergoing apoptosis while ovariectomy mainly reduced the cell proliferating ability. Given the known importance of estrogen for bone growth, one would expect that ovariectomy and high-dose administration of estrogen would have opposite effects. However, the present study indicated that both low and high concentration had a similar effect: a decrease in the chondrocyte number compared with control, suggesting that estrogen has to be maintained within a narrow range for optimal bone growth