Effect of trunk exercise upon lumbar IVD height and vertebral compliance when performed supine with 1 g at the CoM compared to upright in 1 g.

BACKGROUND: Spinal unloading in microgravity is associated with stature increments, back pain, intervertebral disc (IVD) swelling and impaired spinal kinematics. The aim of this study was to determine the effect of lateral stabilization, trunk rotation and isometric abdominal exercise upon lumbar IVD height, and both passive and active vertebral compliance when performed supine on a short-arm human centrifuge (SAHC)-a candidate microgravity countermeasure-with 1 g at the CoM, compared to that generated with equivalent upright exercise in 1 g. METHODS: 12 (8 male) healthy subjects (33.8 ± 7 years, 178.4 ± 8.2 cm, 72.1 ± 9.6 kg) gave written informed consent. Subjects performed three sets of upper body trunk exercises either when standing upright (UPRIGHT), or when being spun on the SAHC. Lumbar IVD height and vertebral compliance (active and passive) were evaluated prior to SAHC (PRE SAHC) and following the first SAHC (POST SPIN 1) and second Spin (POST SPIN 2), in addition to before (PRE UPRIGHT), and after upright trunk exercises (POST UPRIGHT). RESULTS: No significant effect upon IVD height (L2-S1) when performed UPRIGHT or on the SAHC was observed. Trunk muscle exercise induced significant (p < 0.05) reduction of active thoracic vertebral compliance when performed on the SAHC, but not UPRIGHT. However, no effect was observed in the cervical, lumbar or across the entire vertebral column. On passive or active vertebral compliance. CONCLUSION: This study, the first of its kind demonstrates that trunk exercise were feasible and tolerable. Whilst trunk muscle exercise appears to have minor effect upon IVD height, it may be a candidate approach to mitigate-particularly active-vertebral stability on Earth, and in µg via concurrent SAHC. However, significant variability suggests larger studies including optimization of trunk exercise and SAHC prescription with MRI are warranted. TRIAL REGISTRATION: North Rhine ethical committee (Number: 6000223393) and registered on 29/09/2020 in the German Clinical Trials Register (DRKS00021750).

Sodium hypochlorite accident with evaluation by cone beam computed tomography.

AIM: To show the radiographic manifestation of sodium hypochlorite after accidental injection past the apical foramen and into the soft tissues. SUMMARY: A female patient was seen for an emergency visit after suffering a sodium hypochlorite accident at her general dentist's office. The patient was seen within 1 h of the accident and was in pain associated with facial swelling. Radiographs, including a Cone Beam Computed Tomography (CBCT), and photographs were taken. Endodontic emergency treatment was initiated. The patient was reassured and given pain medication and antibiotics. Follow-up visits were scheduled over 6 days when the swelling had resolved. KEY LEARNING POINTS: • Importance of multiple radiographic images during preoperative endodontic evaluation when undertaking endodontic retreatment. • Knowledge of apical anatomy as related to surrounding structures. • Effect of sodium hypochlorite when injected in the soft tissues.

A functional alternative splicing mutation in human tryptophan hydroxylase-2.

The brain serotonergic system has an essential role in the physiological functions of the central nervous system and dysregulation of serotonin (5-HT) homeostasis has been implicated in many neuropsychiatric disorders. The tryptophan hydroxylase-2 (TPH2) gene is the rate-limiting enzyme in brain 5-HT synthesis, and thus is an ideal candidate gene for understanding the role of dysregulation of brain serotonergic homeostasis. Here, we characterized a common, but functional single-nucleotide polymorphism (SNP rs1386493) in the TPH2 gene, which decreases efficiency of normal RNA splicing, resulting in a truncated TPH2 protein (TPH2-TR) by alternative splicing. TPH2-TR, which lacks TPH2 enzyme activity, dominant-negatively affects full-length TPH2 function, causing reduced 5-HT production. The predicted mRNA for TPH2-TR is present in postmortem brain of rs1386493 carriers. The rs13864923 variant does not appear to be overrepresented in either global or multiplex depression cohorts. However, in combination with other gene variants linked to 5-HT homeostasis, this variant may exhibit important epistatic influences.

Influence of the elasticity module of synthetic and natural polymeric tissue substitutes on the mobility of the diaphragm and healing process in a rabbit model.

Mesh implants are frequently used in congenital diaphragmatic hernia. This experimental study aimed to examine the influence of different materials on the diaphragmatic movement over time as well as their mechanical qualities after 4 months. Ultrapro®, Surgisis®, and Proceed® were implanted onto a diaphragmatic defect in growing rabbits. Diaphragmatic mobility was determined at three time points. At 4 months, defect shrinkage and mechanical properties were measured. The break strength decreased for Ultrapro® and Surgisis®, but did not change relevantly for Proceed®. Ultrapro® (32.46 N/cm) and Proceed® (31.75 N/cm) showed a four-fold higher resistance to tearing than Surgisis® (8.31 N/cm). The elasticity of Ultrapro® showed no significant difference compared to Surgisis® (p = 0.75). Proceed®, on the other hand, was more than twice as elastic as Ultrapro® or Surgisis® (p = 0.015). Ultrapro® had a higher spring rate (6.48 N/mm) compared to Surgisis® (3.82 N/mm) or Proceed® (5.23 N/mm). Observing the standardized movement rates of the diaphragm for each mesh group over time the only statistical differences were seen for the Proceed® group. On account of its material qualities Ultrapro® was found to be the most suitable mesh material for demanding locations in our model.

A Community's Awareness and Perceptions of Genomic Medicine.

BACKGROUND: Focus groups were conducted in a mid-sized community to explore community members' awareness and perceptions of genomic medicine and identify effective methods to educate the public about this topic. METHODS: Thirteen focus groups were conducted with a demographically representative pool of 121 participants using a semi-structured interview guide. Transcripts were analyzed through a computer assisted approach with Atlas TI consisting of coding, categorizing, comparing, and contrasting relevant data. RESULTS: Identified categories were organized into 6 main themes, which were similar across the groups and included: a lack of awareness, perceived benefits, concerns about genomic medicine, reasons for poor health related behavior, the potential impact of genetic information on health behavior, and the best ways to educate the community. Common concerns included lack of affordability, unanticipated physical harm, mistrust of the government and researchers, downstream effects like overpopulation, playing God/disturbing the natural order, lack of regulations, loss of privacy, genetic discrimination, and moral dilemmas posed by genetic engineering, cloning, choosing traits, and abortions resulting from genetic information. Participants also discussed ways to educate the community. CONCLUSIONS: While individuals recognized that diseases run in families, personal experience was a driving factor in participants' level of knowledge. Many expressed optimism about genomic medicine. However, the lack of depth in responses and their misconceptions reflect a deficiency of knowledge, which along with their personal, moral, and global concerns could impede acceptance and utilization of genomic medicine. Many community members are receptive to learning more about genomic medicine, and many of their concerns and misconceptions can be addressed through a well designed education strategy.

Maternal fumonisin exposure as a risk factor for neural tube defects.

Fumonisins are mycotoxins produced by the fungus F. verticillioides, a common contaminant of maize (corn) worldwide. Maternal consumption of fumonisin B(1)-contaminated maize during early pregnancy has recently been associated with increased risk for neural tube defects (NTDs) in human populations that rely heavily on maize as a dietary staple. Experimental administration of purified fumonisin to mice early in gestation also results in an increased incidence of NTDs in exposed offspring. Fumonisin inhibits the enzyme ceramide synthase in de novo sphingolipid biosynthesis, resulting in an elevation of free sphingoid bases and depletion of downstream glycosphingolipids. Increased sphingoid base metabolites (i.e., sphinganine-1-phosphate) may perturb signaling cascades involved in embryonic morphogenesis by functioning as ligands for sphingosine-1-P (S1P) receptors, a family of G-protein-coupled receptors that regulate key biological processes such as cell survival/proliferation, differentiation and migration. Fumonisin-induced depletion of glycosphingolipids impairs expression and function of the GPI-anchored folate receptor (Folr1), which may also contribute to adverse pregnancy outcomes. NTDs appear to be multifactorial in origin, involving complex gene-nutrient-environment interactions. Vitamin supplements containing folic acid have been shown to reduce the occurrence of NTDs, and may help protect the developing fetus from environmental teratogens. Fumonisins appear to be an environmental risk factor for birth defects, although other aspects of maternal nutrition and genetics play interactive roles in determining pregnancy outcome. Minimizing exposures to mycotoxins through enhanced agricultural practices, identifying biomarkers of exposure, characterizing mechanisms of toxicity, and improving maternal nutrition are all important strategies for reducing the NTD burden in susceptible human populations.

Novel congenital myopathy locus identified in Native American Indians at 12q13.13-14.1.

BACKGROUND: Native American myopathy (NAM) is an autosomal recessive congenital myopathy first reported in the Lumbee Indian people. Features of NAM include congenital weakness, cleft palate, ptosis, short stature, and susceptibility to malignant hyperthermia provoked by anesthesia. METHOD: We identified five individuals with NAM from the Lumbee population, and hypothesized that these affected individuals have disease alleles shared identical-by-descent inherited from common ancestry. To identify a NAM disease locus, homozygosity mapping methods were employed on a genomewide 10K single-nucleotide polymorphism (SNP) screen. To confirm regions of homozygosity identified in the SNP screen, microsatellite repeat markers were genotyped within those homozygous segments. RESULTS: The SNP data demonstrated five regions of shared homozygosity in individuals with NAM. The additional genotyping data narrowed the region to one common segment of homozygosity spanning D12S398 to rs3842936 mapping to 12q13.13-14.1. Notably, loss of heterozygosity estimates from the SNP data also detected this same 12q region in the affected individuals. CONCLUSION: This study reports the first gene mapping of Native American myopathy (NAM) using single-nucleotide polymorphism-based homozygosity mapping in only a few affected individuals from simplex families and identified a novel NAM locus. Identifying the genetic basis of NAM may suggest new genetic etiologies for other more common conditions such as congenital myopathy and malignant hyperthermia.

Genotype-phenotype study in an FSHD family with a proximal deletion encompassing p13E-11 and D4Z4.

BACKGROUND: In the majority of facioscapulohumeral muscular dystrophy (FSHD) cases, the molecular basis of the disease is due to loss of subtelomeric D4Z4 repeat units at 4q35. Occasionally, an apparent absence of the contracted D4Z4 repeat is associated with FSHD. One explanation for this finding is a deletion in the region proximal to the D4Z4 repeat array that encompasses the p13E-11 (D4F104S1) probe-binding site used in the DNA diagnosis. The frequency of such proximally extended deletions is unknown, and to date, few patients have been described due to the difficulties in the molecular identification of such cases. METHODS: We describe a family (DUK 2531) in which a contracted D4Z4 allele and a large proximal deletion of approximately 75 kb are segregating to 11 individuals. This is the largest deletion identified to date. Family DUK 2531 was initially thought to have normal D4Z4 fragment size and therefore unlinked to the 4q35 region (FSHD1B). RESULTS: Further molecular analysis of DUK 2531 reveals the presence of 10 repeat units (33 kb). The extended deletion includes the probe p13E-11 and B31 binding sites, the inverted repeat D4S2463, and genes FRG2 and TUBB4Q. CONCLUSION: Despite the length of the proximal deletion in this family, the range and severity of the clinical manifestations are typical for the disorder. Because such deletions can lead to misinterpretation in the diagnostic setting, this suggests the need for additional diagnostic tests in facioscapulohumeral muscular dystrophy.

Power calculations for likelihood ratio tests for offspring genotype risks, maternal effects, and parent-of-origin (POO) effects in the presence of missing parental genotypes when unaffected siblings are available.

Genotype-based likelihood-ratio tests (LRT) of association that examine maternal and parent-of-origin effects have been previously developed in the framework of log-linear and conditional logistic regression models. In the situation where parental genotypes are missing, the expectation-maximization (EM) algorithm has been incorporated in the log-linear approach to allow incomplete triads to contribute to the LRT. We present an extension to this model which we call the Combined_LRT that incorporates additional information from the genotypes of unaffected siblings to improve assignment of incompletely typed families to mating type categories, thereby improving inference of missing parental data. Using simulations involving a realistic array of family structures, we demonstrate the validity of the Combined_LRT under the null hypothesis of no association and provide power comparisons under varying levels of missing data and using sibling genotype data. We demonstrate the improved power of the Combined_LRT compared with the family-based association test (FBAT), another widely used association test. Lastly, we apply the Combined_LRT to a candidate gene analysis in Autism families, some of which have missing parental genotypes. We conclude that the proposed log-linear model will be an important tool for future candidate gene studies, for many complex diseases where unaffected siblings can often be ascertained and where epigenetic factors such as imprinting may play a role in disease etiology.

Potential for expanded power in linkage studies using the ALLEGRO and MERLIN software programs.

Multipoint linkage analysis in complex diseases requires the use of fast algorithms that can handle many markers and a large number of moderately sized pedigrees with unknown mode of inheritance. This need has led to the development of several competitive software programs. We recently completed a genomic screen of neural tube defects using GENEHUNTER-PLUS and the more recent ALLEGRO. The ALLEGRO software was found to offer expanded power for linkage studies, particularly for childhood onset diseases like neural tube defects, though the results must be treated with caution.

Whole genomewide linkage screen for neural tube defects reveals regions of interest on chromosomes 7 and 10.

Neural tube defects (NTDs) are the second most common birth defects (1 in 1000 live births) in the world. Periconceptional maternal folate supplementation reduces NTD risk by 50-70%; however, studies of folate related and other developmental genes in humans have failed to definitively identify a major causal gene for NTD. The aetiology of NTDs remains unknown and both genetic and environmental factors are implicated. We present findings from a microsatellite based screen of 44 multiplex pedigrees ascertained through the NTD Collaborative Group. For the linkage analysis, we defined our phenotype narrowly by considering individuals with a lumbosacral level myelomeningocele as affected, then we expanded the phenotype to include all types of NTDs. Two point parametric analyses were performed using VITESSE and HOMOG. Multipoint parametric and nonparametric analyses were performed using ALLEGRO. Initial results identified chromosomes 7 and 10, both with maximum parametric multipoint lod scores (Mlod) >2.0. Chromosome 7 produced the highest score in the 24 cM interval between D7S3056 and D7S3051 (parametric Mlod 2.45; nonparametric Mlod 1.89). Further investigation demonstrated that results on chromosome 7 were being primarily driven by a single large pedigree (parametric Mlod 2.40). When this family was removed from analysis, chromosome 10 was the most interesting region, with a peak Mlod of 2.25 at D10S1731. Based on mouse human synteny, two candidate genes (Meox2, Twist1) were identified on chromosome 7. A review of public databases revealed three biologically plausible candidates (FGFR2, GFRA1, Pax2) on chromosome 10. The results from this screen provide valuable positional data for prioritisation of candidate gene assessment in future studies of NTDs.

The genes encoding for D4Z4 binding proteins HMGB2, YY1, NCL, and MYOD1 are excluded as candidate genes for FSHD1B.

Facioscapulohumeral muscular dystrophy is a disease of skeletal muscle, with symptoms including both facial and shoulder girdle weakness and progression to involve the pelvic girdle and extremities in the majority of cases. For most cases of FSHD, the molecular basis of the disease can be identified as a partial deletion of the D4Z4 repeat array on the end of the long arm of chromosome 4. However, in up to 5% of FSHD families there is no linkage to 4q35. These cases are designated as FSHD1B. Proteins have been identified that bind to the D4Z4 repeats of chromosome 4q35. The genes encoding D4Z4 binding proteins YY1, HMGB2, NCL, and MYOD1 were investigated as candidate genes for FSHD1B. Coding sequences and promoter region were analyzed for HMBG2 and no sequence variations were detected. For YY1, all five exons were analyzed and a polymorphism was detected in both the unaffected and affected populations. In nucleolin (NCL), several SNPs were identified, including a SNP causing the non-synonymous change P515H; however, all polymorphisms either occurred in control samples or were previously reported. A novel polymorphism was also detected in MYOD1, but did not represent a disease-specific variation. These results suggest that HMBG2, YY1, NCL, and MYOD1 are unlikely to represent the genes responsible for FSHD in these families.

Cloning and characterization of an inversion breakpoint at 6q23.3 suggests a role for Map7 in sacral dysgenesis.

Here we report on a male patient with sacral dysgenesis (SD) and constitutional pericentric inversion of chromosome 6 (p11.2;q23.3). SD is a heterogeneous group of congenital anomalies with complex genetic etiology. Previously, a patient with sacral abnormalities and an interstitial deletion of 6q23-->q25 region has been described. We speculated that a susceptibility gene for SD lies in 6q23.3 region (disrupted in both patients), and therefore, cloning of the breakpoint in our patient would lead to the identification of the disrupted gene. We performed FISH analysis followed by Southern blot analysis and inverse PCR to clone the breakpoint. The 6p11.2 breakpoint mapped very close to the centromere, and the 6q23.3 breakpoint localized in the ninth intron of the MAP7 gene. We then evaluated the involvement of MAP7 in SD by further screening of the gene in several patients with a similar phenotype. Two nucleotide changes causing Ile257Asn and Glu571Ala substitutions in the protein, both affecting amino acid residues conserved in the mouse homolog, were identified in two patients. Both changes are either very rare polymorphisms or true mutations, since they were not detected in 167 normal individuals nor found in the SNP database. Therefore, our study suggests MAP7 as a candidate gene for SD. However, we were unable to detect any sacral defects in the MAP7 knockout mice.

Updated investigations of the role of methylenetetrahydrofolate reductase in human neural tube defects.

Folate supplementation appears to reduce the risk for neural tube defects (NTDs). Methylenetetrahydrofolate reductase (MTHFR) is a candidate gene in the folate metabolism pathway that has been extensively studied in different human populations. We examined the risk associated with having the thermolabile variant (TT) of MTHFR in a study of 175 American Caucasians with NTDs and their families. We found a significant association in patients compared with 195 unrelated controls [odds ratio (OR) = 2.13, 95% confidence interval (95% CI) = 1.11-4.09)], but not in mothers (OR = 1.29, 95% CI = 0.622-2.67) or in fathers (OR = 1.45, 95% CI = 0.681-3.09). We found no evidence for unequal transmission from parents to an affected child (p > 0.10). We failed to find a previously reported association for a combined haplotype for MTHFR and cystathionine beta-synthase, except in subjects with NTDs compared with 559 pooled controls (OR = 2.87, 95% CI = 1.03-8.03). We found no evidence for an association for a novel CA-repeat polymorphism identified in a gene closely linked to MTHFR (p > 0.10). Our studies continue to suggest that additional candidate genes other than MTHFR may be responsible for an increased risk to NTD in some American Caucasian families.

Smooth muscle cell phenotypic transition associated with calcification: upregulation of Cbfa1 and downregulation of smooth muscle lineage markers.

Bovine aortic smooth muscle cell (BASMC) cultures undergo mineralization on addition of the organic phosphate donor, beta-glycerophosphate (betaGP). Mineralization is characterized by apatite deposition on collagen fibrils and the presence of matrix vesicles, as has been described in calcified vascular lesions in vivo as well as in bone and teeth. In the present study, we used this model to investigate the molecular mechanisms driving vascular calcification. We found that BASMCs lost their lineage markers, SM22alpha and smooth muscle alpha-actin, within 10 days of being placed under calcifying conditions. Conversely, the cells gained an osteogenic phenotype as indicated by an increase in expression and DNA-binding activity of the transcription factor, core binding factor alpha1 (Cbfa1). Moreover, genes containing the Cbfa1 binding site, OSE2, including osteopontin, osteocalcin, and alkaline phosphatase were elevated. The relevance of these in vitro findings to vascular calcification in vivo was further studied in matrix GLA protein null (MGP(-/-)) mice whose arteries spontaneously calcify. We found that arterial calcification was associated with a similar loss in smooth muscle markers and a gain of osteopontin and Cbfa1 expression. These data demonstrate a novel association of vascular calcification with smooth muscle cell phenotypic transition, in which several osteogenic proteins including osteopontin, osteocalcin, and the bone determining factor Cbfa1 are gained. The findings suggest a positive role for SMCs in promoting vascular calcification.

Additional glomangioma families link to chromosome 1p: no evidence for genetic heterogeneity.

Venous malformations are a common abnormality of the vasculature that may occur sporadically or, more rarely, as an autosomal dominant trait. One familial form of venous malformations has previously been linked to chromosome 9p. Mutations in the gene encoding Tie2, an endothelial specific receptor tyrosine kinase, have been identified in four different families. Glomangiomas are a subtype of venous malformations with glomus cell involvement. These cutaneous lesions can be inherited as an autosomal dominant disease with reduced penetrance and variable expressivity. We present evidence of linkage to chromosome 1p21-1p22 using four new glomangioma families, with a combined maximum two-point lod score of 7.32 at marker D1S2804. Markers D1S2129 and D1S2881 define the 24-cM linkage interval determined by recombination within affected individuals. A recent report also showed linkage of the glomangioma locus to chromosome 1p. A total of 9 families now map to this region, suggesting a decreased likelihood of locus heterogenity in familial glomangiomas. Investigation of candidate genes within the interval should provide new insights into lesion formation in inherited venous malformations.

Clinical Studies in Non-chromosome 4-Linked Facioscapulohumeral Muscular Dystrophy.

OBJECTIVES: To characterize clinically and molecularly a large, non-chromosome 4-linked facioscapulohumeral muscular dystrophy (FSHMD) family. METHODS: Neurological evaluations of affected (N = 55) and at-risk (N = 48) individuals were performed along with selected laboratory analyses, including creatine kinase testing, muscle biopsy, p13E-11 fragment analysis, and cytogenetic studies. Genetic analyses of the scapuloperoneal muscular dystrophy and scapuloperoneal muscular atrophy regions on chromosome 12 were performed using genetic markers flanking the intervals of interest and parametric LOD score analyses. RESULTS: Clinically, the FSHMD in individuals in this family is indistinguishable from that observed in chromosome 4-linked FSHMD. Fragment analysis with p13E-11 showed no small fragment segregating with the family and no evidence for 4:10 translocation or deletion of the p13E-11 binding site. Linkage analysis excluded the loci for autosomal-dominant scapuloperoneal muscular dystrophy and scapuloperoneal muscular atrophy. CONCLUSIONS: This family is clinically similar to patients with the chromosome 4-linked FSHMD. These data support our previous hypothesis of genetic heterogeneity within FSHMD.

Life after the screen: making sense of many P-values.

A multiple analytic approach may be useful for analyzing complex traits since different methods extract both similar and distinct, but complementary pieces of information from genome screen data on extended pedigrees. We examined the usefulness of combining p-values both across methods and across adjacent markers, taking into account the observed correlation structure among these p-values. To this end, we employed the recently proposed truncated product method [Zaykin et al., Genet Epidemiol, in press]. It appears that this approach is helpful for visualizing priority regions for follow-up analysis and reducing the number of false-positive linkage signals.