Evaluation of the prenatal diagnosis of limb reduction deficiencies. EUROSCAN Study Group.

Ultrasound scans in the mid-trimester of pregnancy are now a routine part of antenatal care in most European countries. Using data from registries of congenital anomalies a study was undertaken in Europe. The objective of the study was to evaluate prenatal detection of limb reduction deficiencies (LRD) by routine ultrasonographic examination of the fetus. All LRDs suspected prenatally and all LRDs (including chromosome anomalies) confirmed at birth were identified from 20 Congenital Malformation Registers from the following 12 European countries: Austria, Croatia, Denmark, France, Germany, Italy, Lithuania, Spain, Switzerland, The Netherlands, UK and Ukrainia. These registries are following the same methodology. During the study period (1996-98) there were 709,030 births, and 7,758 cases with congenital malformations including LRDs. If more than one LRD was present the case was coded as complex LRD; 250 cases of LRDs with 63 (25.2%) termination of pregnancies were identified including 138 cases with isolated LRD, 112 with associated malformations, 16 with chromosomal anomalies and 38 non chromosomal recognized syndromes. The prenatal detection rate of isolated LRD was 24.6% (34 out of 138 cases) compared with 49.1% for associated malformations (55 out of 112; p<0.01). The prenatal detection of isolated terminal transverse LRD was 22.7% (22 out of 97), 50% (3 out of 6) for proximal intercalary LRD, 8.3% (1 out of 12) for longitudinal LRD and 0 for split hand/foot; for multipli-malformed children with LRD those percentages were 46.1% (30 out of 65), 66.6% (6 out of 9), 57.1% (8 out of 14) and 0 (0 out of 2), respectively. The prenatal detection rate of LRDs varied in relation with the ultrasound screening policies from 20.0% to 64.0% in countries with at least one routine fetal scan.

Angiographic demonstration of neoangiogenesis after intra-arterial infusion of autologous bone marrow mononuclear cells in diabetic patients with critical limb ischemia.

Critical limb ischemia in diabetic patients is associated with high rates of morbidity and mortality. Suboptimal responses to the available medical and surgical treatments are common in these patients, who also demonstrate limited vascular homeostasis. Neovasculogenesis induced by stem cell therapy could be a useful approach for these patients. Neovasculogenesis and clinical improvement were compared at baseline and at 3 and 12 months after autologous bone marrow-derived mononuclear cell (BMMNC) transplantation in diabetic patients with peripheral artery disease. We conducted a prospective study to evaluate the safety and efficacy of intra-arterial administration of autologous BMMNCs (100-400 × 10(6) cells) in 20 diabetic patients with severe below-the-knee arterial ischemia. Although the time course of clinical effects differed among patients, after 12 months of follow-up all patients presented a notable improvement in the Rutherford-Becker classification, the University of Texas diabetic wound scales, and the Ankle-Brachial Index in the target limb. The clinical outcome was consistent with neovasculogenesis, which was assessed at 3 months by digital subtraction angiography and quantified by MetaMorph software. Unfortunately, local cell therapy in the target limb had no beneficial effect on the high mortality rate in these patients. In diabetic patients with critical limb ischemia, intra-arterial perfusion of BMMNCs is a safe procedure that generates a significant increase in the vascular network in ischemic areas and promotes remarkable clinical improvement.

Spatio-temporal micromeasurements of the oxygen uptake in the developing chick embryo.

A method has been developed for the determination of the oxygen uptake of small areas (0.01 mm2) in an entire chick embryo cultured in vitro under defined metabolic conditions. It is based on the recordings of the spectral changes of the hemoglobin used as oxygen source for the respiring tissue (Barzu and Borza, 1967). Rapid scanning of the hemoglobin absorbance over the preparation allows a comparison of the O2 uptake of various regions. Values of the order of 10(-2) 1 O2 . min-2 are measured in less than 10 sec with a spatial resolution of 100 micron. The differentiation of embryonic tissue is not disturbed by the measurements. The O2 diffusion in the media and in the tissue has been analyzed by digital simulation. The O2 uptake of the Hensen's node was measured from embryos starting at the stage of definitive primitive streak (stage 4) up to the stage of 10 somites. It increases from 0.6 to 1.1 nl . h-1 with a marked acceleration between stages 4 and 5. The values corrected for the protein content of the Hensen's node at stage 4, 5, 6 and 8 are 32, 30 and 28 microliter . mg-1 . h-1 respectively. The first scanning results show different patterns of the O2 utake at the level of the Hensen's node and of the neural plate. At stage 6-7, the corrected O2 uptake is 30 microliter . mg-1 . h-1 for . the former and 43 microliter . mg-1 . h-1 for the latter.

Simultaneous Doppelgänger and limb amputation impression in right frontal opercular stroke.

A case is described of a patient who presented almost simultaneously the impression that his left arm was amputated and the feeling of the presence of his invisible Doppelgänger. While these body scheme disorders have both been described after (right) parietal lesions, a right frontal opercular ischaemic stroke was found in the neurological work up. Diffusion tensor imaging showed that the stroke involved the ventral bundle of the superior longitudinal fasciculus that connects the parietal to the frontal lobe. The unusual clinical presentation of this frontal lesion may have been due to a 'diaschisis'-like phenomenon via the superior longitudinal fasciculus.

Fibronectin distribution in the chick embryo during formation of the blastula.

The distribution of the fibronectin-rich extracellular matrix (ECM) in the chick embryo during formation of the blastula has been evaluated semiquantitatively using an electron microscopical immunogold staining technique. During the first 10 h of postlaying development, fibronectin was found in both embryonic area pellucida and extra-embryonic area opaca of the blastoderm. In the area pellucida, the fibronectin was (1) associated with the basal lamina of the epiblast, (2) present between epiblastic and hypoblastic cells and (3) occasionally internalized in hypoblastic cells. Along the embryonic axis, a transient and high density of ECM was associated with the front of the anteriorly and rapidly expanding hypoblast. Very high density of fibronectin was observed in the marginal zone of the area pellucida, where the epiblastic and deeper cell layers show contacts and intense re-arrangements. In the area opaca, fibronectin was at first found only sporadically between contacting cells, but its density increased steadily and markedly during the first day of development. These rapid and significant changes in the regional distribution of fibronectin-rich ECM are discussed with respect to the early morphogenesis of the chick embryo.

CO2 production of the chick embryo during the first day of post-laying development.

The carbon dioxide production of the chick embryo cultured in vitro has been determined during the first 24 h of post-laying development using a non-invasive conductometric microtechnique. The mean CO2 production of the whole blastoderm (1) increased from 16 nmol/h at laying to 231 nmol/h at early neurulation, (2) became dependent on exogenous glucose and (3) was closely linked to mechanical tension generated in the blastoderm (loosening from vitelline membrane resulted in a decrease of 56%). In our experimental conditions, no significant influence of carbonic anhydrase on the CO2 production has been detected. The value of the respiratory exchange ratio varied from about 3 at pregastrular stages to 1 at neurula stage and CO2 was produced transiently in presence of antimycin A. Such results indicate that the source of CO2 is not exclusively mitochondrial and that the relative proportions of mitochondrial and non-mitochondrial CO2 productions might vary significantly throughout the early development. Our findings confirm that the metabolism of the chick embryo becomes more and more oxidative from laying onwards and suggest that the modifications of metabolism observed during the studied period of development could be associated with functional differentiation.

Oxidative and glycogenolytic cCapacities within the developing chick heart.

Cardiac morphogenesis and function are known to depend on both aerobic and anaerobic energy-producing pathways. However, the relative contribution of mitochondrial oxidation and glycogenolysis, as well as the determining factors of oxygen demand in the distinct chambers of the embryonic heart, remains to be investigated. Spontaneously beating hearts isolated from stage 11, 20, and 24HH chick embryos were maintained in vitro under controlled metabolic conditions. O(2) uptake and glycogenolytic rate were determined in atrium, ventricle, and conotruncus in the absence or presence of glucose. Oxidative capacity ranged from 0.2 to 0.5 nmol O(2)/(h.microg protein), did not depend on exogenous glucose, and was the highest in atria at stage 20HH. However, the highest reserves of oxidative capacity, assessed by mitochondrial uncoupling, were found at the youngest stage and in conotruncus, representing 75 to 130% of the control values. At stage 24HH, glycogenolysis in glucose-free medium was 0.22, 0.17, and 0.04 nmol glucose U(h.microg protein) in atrium, ventricle, and conotruncus, respectively. Mechanical loading of the ventricle increased its oxidative capacity by 62% without altering glycogenolysis or lactate production. Blockade of glycolysis by iodoacetate suppressed lactate production but modified neither O(2) nor glycogen consumption in substrate-free medium. These findings indicate that atrium is the cardiac chamber that best utilizes its oxidative and glycogenolytic capacities and that ventricular wall stretch represents an early and major determinant of the O(2) uptake. Moreover, the fact that O(2) and glycogen consumptions were not affected by inhibition of glyceraldehyde-3-phosphate dehydrogenase provides indirect evidence for an active glycerol-phosphate shuttle in the embryonic cardiomyocytes.

A dystroglycan mutation associated with limb-girdle muscular dystrophy.

Dystroglycan, which serves as a major extracellular matrix receptor in muscle and the central nervous system, requires extensive O-glycosylation to function. We identified a dystroglycan missense mutation (Thr192→Met) in a woman with limb-girdle muscular dystrophy and cognitive impairment. A mouse model harboring this mutation recapitulates the immunohistochemical and neuromuscular abnormalities observed in the patient. In vitro and in vivo studies showed that the mutation impairs the receptor function of dystroglycan in skeletal muscle and brain by inhibiting the post-translational modification, mediated by the glycosyltransferase LARGE, of the phosphorylated O-mannosyl glycans on α-dystroglycan that is required for high-affinity binding to laminin.

A new way of quantifying dominant upper-limb mobility in healthy and painful shoulders

Introduction: Quantification of daily upper-limb activity is determinant in the evaluation of shoulder surgery. For a number of shoulder diseases, roblems in performing daily activities have been expressed in terms of upper-limb usage. Althought many instruments measure upper-limb movements, there is no accepted standard or widely used objective measure and no device to differenciate left or right shoulder usage. We present an objective method to measure the mobility and quantify the usage of dominant and healthy or painfull shoulder movement during daily life. Methods: 12 patients with unilateral pathological shoulder (rotator cuff disease) are compared to 18 control subjects (10 right and 8 left handed). Both SST and DASH questionnaires were completed by each one. Three inertial miniature modules including triaxial gyroscopes and accelerometers were fixed on the dorsal side of both humerus, and on the thorax. An ambulatory datalogger have recorded the signals during one day. Results: We observed that right handed healthy subjects used 18% and 26% more their dominant shoulder during respectively stand and sit postures while left handed subjects used 8% and 18% more their left side. In walking periods, shoulder mobility was quite alike for both sides. Patients affected on their dominant arm (PD group) mostly used their non-dominant side (respectively 5% and 9% during stand and sit). For the patients affected on their non-dominant shoulder (PND group), this difference is respectively 28% and 26%. Moreover, we can note that, during walking periods, a difference can be observed (on the contrary to controls). Patients used 13% and 15% more their nonpathologic side respectively for PD and PND groups. Conclusion: Inertial sensors, during long-term ambulatory monitoring of upper limbs, can quantify the difference between dominant and nondominant sides. Patients used more their non affected shoulder during daily life. For the PD group, the difference with control can be shown during walking. These results are very encouraging for future evaluation of patients with shoulder injuries since it can provide an objective evaluation of the shoulder mobility and of the treatment outcome during daily life.

Preferential synthesis of prostaglandin D2 by neurons and prostaglandin E2 by fibroblasts and nonneuronal cells in chick dorsal root ganglia.

To determine the type and the relative amount of prostaglandins (PGs) synthesized by various neural tissues, homogenates of meninges, dorsal root ganglia (DRG) capsules, decapsulated DRG, and unsheathed sciatic nerves were incubated with [1-14C]arachidonic acid. Homogenates of cultured cells (meningeal cells, fibroblasts, and nonneuronal or neuronal DRG cells) were used to specify the cells producing particular PGs. The highest synthetic capacity was found in fibroblast-rich tissues (meninges and DRG capsules) and in cultures of meningeal cells or fibroblasts. Two major cyclooxygenase products were formed: [14C]PGE2 and an unusual 14C-labeled compound, Y. The accumulation of compound Y, corresponding probably to 15-hydroperoxy PGE2, was completely impaired by addition of exogenous GSH, which conversely enhanced the synthesis of [14C]PGE2 and promoted the formation of [14C]PGD2. In contrast, decapsulated DRG or unsheathed sciatic nerves displayed a 10-20 times lower capacity to synthesize PGs than fibroblast-rich tissues and produced mainly [14C]PGE2 and [14C]PGD2. In this case, [14C]PGE2 or [14C]PGD2 synthesis was neither enhanced nor promoted by addition of exogenous GSH. Neuron-enriched DRG cell cultures allowed us to specify that [14C]PGD2 is the major prostanoid produced by primary sensory neurons as compared with nonneuronal DRG cells. Because PGD2 synthesis in DRG and more specifically in DRG neurons does not depend on exogenous GSH and differs from PGD2 synthesis in fibroblast-rich tissues, it is concluded that at least two distinct enzymatic processes contribute to PGD2 formation in the nervous system.(ABSTRACT TRUNCATED AT 250 WORDS)

Substance P-like-immunoreactive sensory neurons in dorsal root ganglia of the chick embryo: ontogenesis and influence of peripheral targets.

The expression of substance P (SP) was studied in sensory neurons of developing chick lumbosacral dorsal root ganglia (DRG) by using a mixture of periodic acid, lysine and paraformaldehyde as fixative and a monoclonal antibody for SP-like immunostaining. The first SP-like-immunoreactive DRG cells appeared first at E5, then rapidly increased in number to reach a peak (88% of ganglion cells) at E8, and finally declined (59% at E12, 51% after hatching). The fall of the SP-like-positive DRG cells resulted from two concomitant events affecting a subset of small B-neurons: a loss of neuronal SP-like immunoreactivity and cell death. After one hindlimb resection at an early (E6) or late (E12) stage of development (that is before or after establishment of peripheral connections), the DRG were examined 6 days later. In both cases, a drastic neuronal death occurred in the ispilateral DRG. However, the resection at E6 did not change the percentage of SP-like-positive neurons, while the resection at E12 severely reduced the proportion of SP-like-immunoreactive DRG cells (25%). In conclusion, connections established between DRG and peripheral target tissues not only promote the survival of sensory neurons, but also control the maintenance of SP-like-expression. Factors issued from innervated targets such as NGF would support the survival of SP-expressing DRG cells and enhance their SP content while other factors present in skeletal muscle or skin would hinder SP expression and therefore lower SP levels in a subset of primary sensory neurons.