Comparison of radiograph-based texture analysis and bone mineral density with three-dimensional microarchitecture of trabecular bone.

PURPOSE: Hip fracture is a serious health problem and textural methods are being developed to assess bone quality. The authors aimed to perform textural analysis at femur on high-resolution digital radiographs compared to three-dimensional (3D) microarchitecture comparatively to bone mineral density. METHODS: Sixteen cadaveric femurs were imaged with an x-ray device using a C-MOS sensor. One 17 mm square region of interest (ROI) was selected in the femoral head (FH) and one in the great trochanter (GT). Two-dimensional (2D) textural features from the co-occurrence matrices were extracted. Site-matched measurements of bone mineral density were performed. Inside each ROI, a 16 mm diameter core was extracted. Apparent density (Dapp) and bone volume proportion (BV/TV(Arch)) were measured from a defatted bone core using Archimedes' principle. Microcomputed tomography images of the entire length of the core were obtained (Skyscan 1072) at 19.8 microm of resolution and usual 3D morphometric parameters were computed on the binary volume after calibration from BV/TV(Arch). Then, bone surface/bone volume, trabecular thickness, trabecular separation, and trabecular number were obtained by direct methods without model assumption and the structure model index was calculated. RESULTS: In univariate analysis, the correlation coefficients between 2D textural features and 3D morphological parameters reached 0.83 at the FH and 0.79 at the GT. In multivariate canonical correlation analysis, coefficients of the first component reached 0.95 at the FH and 0.88 at the GT. CONCLUSIONS: Digital radiographs, widely available and economically viable, are an alternative method for evaluating bone microarchitectural structure.

Laws' masks descriptors applied to bone texture analysis: an innovative and discriminant tool in osteoporosis.

OBJECTIVE: The objective of this study was to explore Laws' masks analysis to describe structural variations of trabecular bone due to osteoporosis on high-resolution digital radiographs and to check its dependence on the spatial resolution. Laws' masks are well established as one of the best methods for texture analysis in image processing and are used in various applications, but not in bone tissue characterisation. This method is based on masks that aim to filter the images. From each mask, five classical statistical parameters can be calculated. MATERIALS AND METHODS: The study was performed on 182 healthy postmenopausal women with no fractures and 114 age-matched women with fractures [26 hip fractures (HFs), 29 vertebrae fractures (VFs), 29 wrist fractures (WFs) and 30 other fractures (OFs)]. For all subjects radiographs were obtained of the calcaneus with a new high-resolution X-ray device with direct digitisation (BMA, D3A, France). The lumbar spine, femoral neck, and total hip bone mineral density (BMD) were assessed by dual-energy X-ray absorptiometry. RESULTS: In terms of reproducibility, the best results were obtained with the TRE5E5 mask, especially for three parameters: "mean", "standard deviation" and "entropy" with, respectively, in vivo mid-term root mean square average coefficient of variation (RMSCV)%= 1.79, 4.24 and 2.05. The "mean" and "entropy" parameters had a better reproducibility but "standard deviation" showed a better discriminant power. Thus, for univariate analysis, the difference between subjects with fractures and controls was significant (P<10(-3)) and significant for each fracture group independently (P<10(-4) for HF, P=0.025 for VF and P< 10(-3) for OF). After multivariate analysis with adjustment for age and total hip BMD, the difference concerning the "standard deviation" parameter remained statistically significant between the control group and the HF and VF groups (P<5 x 10(-5), and P=0.04, respectively). No significant correlation between these Laws' masks parameters and BMD was obtained. In addition, this study showed the dependence of Laws' masks parameters on image resolution, which confirms the necessity to perform Laws' textural measurement on high-resolution images. CONCLUSION: The reproducibility and discriminant power of the Laws' masks analysis has been demonstrated on bone images; thus, this method constitutes a promising routine technique for the determination of osteoporosis fracture risk from radiographs.

C21orf5, a human candidate gene for brain abnormalities and mental retardation in Down syndrome.

Mental retardation represents the more invalidating pathological aspect of trisomy 21 and has a hard impact on public health. The dosage imbalance of chromosome 21 genes could be the cause of neurological alterations and mental retardation seen in Down syndrome. We studied C21orf5 that we have demonstrated to be overexpressed in Down syndrome tissues, as a candidate gene for trisomy 21. A new optical technology (Rachidi et al., 2000) was used to compare signal intensity and cell density in presumptive embryonic brain compartments, at their boundaries and in higher specialized brain centres during fetal lifespan. We showed a developmentally regulated transcriptional activity of C21orf5 and a regional and cellular specific distribution of gene transcripts during human embryonic and fetal development. A wide but differential expression was detected in the nervous system during embryogenesis with a relatively lower level in the forebrain than in the midbrain and hindbrain and the highest transcription intensity in the future cerebellum. This developmentally regulated expression is maintained during post-embryogenesis and evolves selectively in fetal cerebral, hippocampal and cerebellar areas. Differential and cellular specificity were detected in hippocampus with higher C21orf5 mRNA level in the pyramidal cells compared to granular cells of the dentate gyrus. The expression pattern detected in cortical and cerebellar structures correlates well to the altered cortical lamination and to the lower size of the cerebellum observed in Down syndrome patients. In addition, the patterned differential expression detected in the medial temporal-lobe system, including hippocampal formation and perirhinal cortex, working as control centres of the memory circuits and involved in cognitive processes and memory storage, also corresponds to abnormal brain regions seen in Down syndrome patients. The C21orf5 selective expression in the key brain structures for learning and memory suggests that C21orf5 overexpression could participate in mental retardation pathogenesis in Down syndrome patients.

[Complication of tracheal intubation: severe cervical cellulitis]. / Complication de l'intubation trachéale: cellulite cervicale grave.

A 40-year-old man, victim of a traffic accident has been hospitalized for a severe head trauma. His trachea has been intubated under general anaesthesia with an 8.0 mm ID tube (Vygon). The cerebral scan revealed a surgical subdural haematoma. In the postoperative period, the patient was admitted in surgical intensive care, under sedation and mechanically ventilation. At day 12 the patient developed a cervical cellulitis complicated of a septic shock. The cervical scan showed an important dilatation of the trachea in the site of the tube cuff. The surgical exploration discovered a complete destruction of the anterior face of de trachea on several centimetres of height. The patient died 24 hours later by multiple organ failure.

Virus-like particle formation in Drosophila melanogaster germ cells suggests a complex translational regulation of the retrotransposon cycle and new mechanisms inhibiting transposition.

Transposition of 1731, a Drosophila melanogaster LTR retrotransposon, was investigated in reproductive organs by RNA, protein and VLP distribution during its life cycle. We detected 1731 transcription in oogonia but not in spermatogonia; in all cells during oogenesis but only in primary spermatocytes; and in ovarian cytoplasm but both in nuclei and cytoplasm of primary spermatocytes. By confocal scanning, we showed that whereas Gag protein appeared in all cytoplasms during oogenesis, in testes Gag detection began in late premeiotic primary spermatocytes and increased in elongating spermatids suggesting distinct mechanisms of 1731 transcription and translation regulation. By electron microscopy, we did not detect 1731 VLPs in ovaries, suggesting a complex post-translational control blocking VLP assembly and transposition. Interestingly, in testes we discovered VLP aggregates in cystic cytoplasm of maturing partially individualized spermatids. In testes, we observed two delays in 1731 product expressions, suggesting a complex temporal control mechanism. Transcriptional/translational delay may be determined by accumulation of 1731 RNAs in primary spermatocyte nuclei. Translational/VLP assembly delay may be determined by post-transductional mechanisms controlling +1 frameshift and Pol-protein degradation. Our results indicated two differential mechanisms inhibiting 1731 transposition in Drosophila melanogaster ovaries and testes. In addition, we proposed a new mechanism for transposition control at the cell cycle level.

The CASK/Lin-2 Drosophila homologue, Camguk, could play a role in epithelial patterning and in neuronal targeting.

Drosophila Camguk (Cmg) is a member of the CAMGUK subfamily of the MAGUK family of proteins which are localized at cell junction and other plasma membrane specialized regions, from worms to mammals. The protein structure of Cmg, as the other CAMGUK proteins, is characterized by only one PDZ domain and an additional CaM kinase domain, similar to CaMKII. While the mammalian ortholog CASKs play an important role in synaptic protein targeting and in synaptic plasticity, the Drosophila Cmg role is unknown. To study its potential role, we reported a detailed analysis of mRNA distribution of the Drosophila cmg gene at cellular and developmental level, during embryonic, larval, pupal and adult stages. The transient cmg transcription in midgut and Malpighian tubules may suggest a potential function in cell junction formation and in epithelial tissue patterning. Interestingly, cmg transcription increases substantially during embryonic neuroblast proliferation, becoming predominant in the developing central nervous system (CNS) during embryonic and postembryonic development stages and in the mature brain. In addition, a high transcriptional level was detected in the eye imaginal discs and in the adult retina, demonstrating a specific and continuous expression of cmg in neuroblasts and photoreceptor neurons, from the onset of cytodifferentiation. Our findings suggest that Cmg could play a potential role in transmembrane protein targeting, particularly in synapses. These observations suggest the existence of a common highly conserved mechanism involved in forming and maintaining proper synaptic protein targeting, which are fundamental features of synaptic plasticity, learning and memory. Through its function, the CaM kinase domain-containing Cmg may be involved in signal transduction cascade. Its potential relation to Calmodulin and CaMKII is discussed.

Regional and cellular specificity of the expression of TPRD, the tetratricopeptide Down syndrome gene, during human embryonic development.

The TPRD gene (tetratricopeptide (TPR) containing Down syndrome gene) is one of the candidate genes in the Down syndrome chromosomal region-1. Duplication of this gene may be the cause of major phenotypic features of Down syndrome. Here we show that the TPRD expression is developmentally regulated during human embryogenesis. At the earliest stages of development (Carnegie 8-12) TPRD expression is ubiquitous. At later developmental stages (Carnegie stages 14, 16 and 18), it becomes restricted to the nervous system, as is the case for the mtprd gene during mouse development. We extended our analysis of TPRD expression during fetal development of the human nervous system (13, 22 and 24 weeks). A new oblique illumination technique was used to compare signal intensity and cell density. Some regions of the nervous system such as the external cortical layers of the brain, and the inner neuroblastic layer of the eye, strongly express the TPRD gene.

Characterization of a novel gene, C21orf6, mapping to a critical region of chromosome 21q22.1 involved in the monosomy 21 phenotype and of its murine ortholog, orf5.

Phenotypic and molecular analyses of patients with partial chromosome 21 monosomy enabled us to define a region, spanning 2.4 Mb between D21S190 and D21S226, associated with arthrogryposis, mental retardation, hypertonia, and several facial anomalies. The markers of the region were used to screen a total human PAC library (Ioannou, RZPD). We isolated 57 PACs, which formed primary contigs. EST clusters (UNIGENE collection) located in a 6-Mb interval, between D21S260 and D21S263, were mapped in individual bacterial clones. We mapped the WI-17843 cluster to the PAC clone J12100, which contains the two anchor markers LB10T and LA329. The open reading frame extends over 960 bp, with three putative start codons. The 1695-bp cDNA containing a polyadenylation signal should correspond to the full-length cDNA. From the genomic sequence, we deduced that the gene contained five exons and that there was a putative promoter sequence upstream from exon 1. In silico screening of DNA databases revealed similarity with a murine EST. The corresponding cDNA (1757 bp) sequence was very similar (>85%) to the human cDNA and had an open reading frame of 876 nucleotides. Somatic hybrid mapping localized the cDNA to mouse chromosome 16. EST analyses and RT-PCR indicated that the third exon in the human gene (exon 2 in the mouse) undergoes alternative splicing. Northern blot hybridization showed that the gene was ubiquitously expressed in humans and mice. The longest mouse clone was used to generate riboprobes, which were hybridized to murine embryos at stages E-9.5, E-10.5, E-12.5, E-13.5, and E-14.5-15, to study the pattern of expression during development. Ubiquitous labeling was observed, with strong signals restricted to limited areas of the telencephalon, the mesencephalon, and the interrhombomeric regions in the central nervous system, and other regions of the body such as the limb buds, branchial arches, and somites.

Developmentally regulated expression of mtprd, the murine ortholog of tprd, a gene from the Down syndrome chromosomal region 1.

The gene tprd, which contains three tetratricopeptide domains, has been recently localized in the Down syndrome (DS) chromosomal region 1. We have cloned a cDNA encoding part of the murine ortholog of tprd and used it to characterize the expression pattern of this gene during development and at the adult stage. At E8.5 the expression is uniform. In the later stages of embryogenesis, although expression remains ubiquitous, a pattern of tissues with particularly high expression develops: the strong expression is restricted to non proliferating zones of the nervous system such as the external layer of the cortex, the spinal cord, the cranial and root ganglia and the nerves. In the brain of adult mouse the strongest signals are observed in layers II-III and V-VI of the cortex, in the hippocampus and in the cerebellum, which correspond to the abnormal brain regions seen in DS patients.

Dynamic expression pattern of Ca(2+)/calmodulin-dependent protein kinase II gene in the central nervous system of Drosophila throughout development.

Calcium/calmodulin-dependent protein kinase II (CaM KII) is thought to be involved in the majority of the neuronal functions mediated by intracellular free Ca(2+), and has been implicated in long-term potentiation, learning, and memory. In this work, we have examined in detail the RNA expression pattern for the Drosophila CaM KII gene by in situ hybridization, during embryonic, larval, pupal, and adult stages. Our results indicate that expression of CaM KII was homogeneous in early embryos, but that during development the gene transcription rapidly became restricted to neuroblasts and their progeny in the nervous system. This predominant expression in the nervous system is maintained during late embryogenesis and post-embryonic development. A signal compartmentalization appeared in the larval central nervous system, where the CaM KII expression became progressively concentrated in the anterior ganglia. In the adult brain, a specific expression was more abundant in a subset of neurons around the central brain, particularly the mushroom bodies and the central complex, structures that play an important role in learning and memory.

Expression of the mnb (dyrk) protein in adult and embryonic mouse tissues.

Mnb is a human homologue of the Drosophila minibrain gene which encodes a serine/threonine protein kinase that is required in distinct neuroblast proliferation centers during postembryonic neurogenesis. The high degree of homology of the human gene to the murine gene (dyrk) allowed us to use a human polyclonal anti-mnb antibody to study the expression pattern of the protein in adult and embryonic mouse tissues. Western blot analysis and immunohistochemical methods were used to define the detailed distribution of mnb in adult brain and 17 days mouse embryos. The results show a high expression in the cerebral cortex, the cerebellum, the hippocampus which is in accordance with previous reports of in situ hybridization studies using mRNA probes but also a very strong expression in the epithelial layers of the skin, the retina, the tongue, the intestine and the kidney which has not been described before. Since epithelial cells are highly mitotic cells and since mnb shares sequence similarities with the cdk kinases involved in the regulation of cell division, this result may indicate a important role of mnb in the cell cycle control.

Developmental expression analysis of the 1731 retrotransposon reveals an enhancement of Gag-Pol frameshifting in males of Drosophila melanogaster.

Extensive analyses of Drosophila melanogaster retrotransposon transcriptions in cultured cells or during development have been reported, but little is known about their translation during the development of the fly. Analysis of the translational products of the 1731 Drosophila melanogaster retrotransposon in Kc Drosophila cultured cells has been reported, showing the existence of primary products (Gag and Pol) and of processed polypeptides of various sizes. Study of 1731 retrotransposon expression at both levels of transcription and translation during the development of Drosophila melanogaster, is presented. 1731 transcripts were detected by in situ hybridization and 1731 proteins were detected by immunostaining and immunoblotting in embryos and in adult gonads. 1731 transcripts and proteins were detected in the mesoderm and central nervous system during embryonic development, in nurse cells and follicle cells in adult ovaries and in primary spermatocytes in adult testes. Moreover, Western blot analysis of the 1731 proteins with anti-Gag or anti-Pol antibodies in gonads revealed that the 1731 mRNA could be translated differentially according to the expressing tissue: essentially, ovarian translation and/or processing of 1731 products is different from that operating in testes, where the Gag-Pol fusion polyprotein is the most prominent product. Our results indicate that expression of the 1731 mobile element is regulated not only at the transcriptional level but also at the translational level, and that this regulation is different in the two sexes.

A new gene encoding a putative transcription factor regulated by the Drosophila circadian clock.

Circadian rhythms of locomotor activity and eclosion in Drosophila depend upon the reciprocal autoregulation of the period (per) and timeless (tim) genes. As part of this regulatory loop, per and tim mRNA levels oscillate in a circadian fashion. Other cycling transcripts may participate in this central pacemaker mechanism or represent outputs of the clock. In this paper, we report the isolation of Crg-1, a new circadianly regulated gene. Like per and tim transcript levels, Crg-1 transcript levels oscillate with a 24 h period in light:dark (LD) conditions, with a maximal abundance at the beginning of the night. These oscillations persist in complete darkness and depend upon per and tim proteins. The putative CRG-1 proteins show some sequence similarity with the DNA-binding domain of the HNF3/fork head family of transcription factors. In the adult head, in situ hybridization analysis reveals that per and Crg-1 have similar expression patterns in the eyes and optic lobes.

Genetical analysis of visual system disorganizer (vid), a new gene involved in normal development of eye and optic lobe of the brain in Drosophila melanogaster.

A neuroanatomical screening of a collection of P-element mutagenized flies has been carried out with the aim of finding new mutants affecting the optic lobe of the adult brain in Drosophila melanogaster. We have identified a new gene that is involved in the development of the adult axon array in the optic ganglia and in the ommatidia assembly. We have named this locus visual system disorganizer (vid). Reversional mutagenesis demonstrated that the vid mutant was the result of a P-element insertion in the Drosophila genome and allowed us to generate independent alleles, some of which resulted in semilethality, like the vid original mutant, while the others were completely lethal. A genetic somatic mosaic analysis indicated that the vid gene is required in the eye for its normal development by inductive effects. This analysis also suggests an inductive effect of the vid gene on the distal portion of the optic lobe, particularly the lamina and the first optic chiasma. Moreover, the absence of mutant phenotype in the proximal region of the optic ganglia, including the medulla, the second optic chiasma, and the lobula complex underlying mosaic eyes, is suggestive of an autonomously acting mechanism of the vid gene in the optic lobe. The complete or partial lethality generated by different mutations at the vid locus suggests that this gene's role may not be limited to the visual system, but may also affect a vital function during Drosophila development.

Analysis of period circadian expression in the Drosophila head by in situ hybridization.

The expression of the period (per) gene of Drosophila melanogaster has been studied by in situ hybridization in the adult's head, where it is required for the fly to exhibit behavioral circadian rhythms. We have used non-radioactive in situ hybridization to obtain a high sensitivity and specificity on head sections, with single cell resolution. Consistent with previous per protein- or per reporter gene-expression, per-expressing cells were detected in the optic lobes and the central brain, as well as in the head sensory organs: eyes, ocelli, maxillary palps and proboscis. In the brain and the eyes, circadian fluctuations of the per mRNA abundance were observed in different per expressing cells.

[Some practical problems arising from the use of the PGE/ERAD method in Morocco]. / Quelques problemes pratiques souleves l'occasion de l'application de la methode PGE/ERAD au Maroc.

PIP: This paper describes the problems arising from the multiround survey designed to study fertility and mortality undertaken in Morocco between July, 1972 and June, 1973. 84,000 people were in the sample; 36,000 from urban centers. Each sampling unit was broken up into 1500 persons which represents 300 households. It was found that the standard error increases when the size of the sampling unit diminishes. This was true both for the number of births and the number of deaths. By taking into account the cost, the weekly working time of an enumerator, the supervision and the distance within a sampling unit the author concludes that small sampling units of about 1,000 persons are more suitable for this method. Part time "resident" enumerators and full time "external" enumerators were employed in the survey. Comparison between the 2 categories shows that in order to get good registration of demographic events, the external enumerators are better off for registering the births in the urban areas and the deaths in rural areas. Resident enumerators did better at births registration in rural areas and deaths in urban areas. To insure that all households in a sampling unit are under control, each household, inhabited or uninhabited, should be numbered, and the interval between visits of the enumerator should not exceed 3 months. Strict control and permanent supervision of the enumerators is necessary to make certain that all households are visited and their members listed. In enumerating events at health centers, only those belonging to the housholds of the sampling units were recorded. Suggestions for better control include choosing supervisors from among the best enumerators with field experience and daily visits to enumerators to check questionnaires. Migration is not suitable for study using the multiple round survey because the sampling units need to be larger. Difficulties occur when comparison is made between results of different visits. For Morocco the comparison between the survey and the registration show that 71% of births were recorded during the registration against 74% during the survey, in both rural and urban areas. The recorded percentage of deaths was 62% for the registration, 57% for the survey.^ieng