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1.
Mol Ecol Resour ; 2021 Jan 27.
Artigo em Inglês | MEDLINE | ID: mdl-33503304

RESUMO

The Southern catfish (Silurus meridionalis) is a nocturnal and benthic freshwater fish endemic to the Yangtze River and its tributaries. In this study, we constructed a chromosome-level draft genome of S. meridionalis using 69.7 Gb Nanopore long reads and 49.5 Gb Illumina short reads. The genome assembly was 741.2 Mb in size with a contig N50 of 13.19 Mb. An additional 116.4 Gb of Bionano and 77.4 Gb of Hi-C data were applied to assemble contigs into scaffolds and further into 29 chromosomes, resulting in a 738.9 Mb genome with a scaffold N50 of 28.04 Mb. A total of 22,965 protein-coding genes were predicted from the genome with 22,519 (98.06%) genes functionally annotated. Comparative genomic and transcriptomic analyses revealed a rod-dominated visual system which was responsible for scotopic vision. The absence of cone opsins SWS1 and SWS2 resulted in the lack of UV and blue violet sensitivity. Mutations at key amino acid sites of RH1.1, RH1.2 and RH2 resulted in spectral tuning good for dim light vision and narrow color vision. A higher expression level of rod phototransduction genes than that of cone genes and higher rod-to-cone ratio led to higher optical sensitivity under dim light conditions. In addition, analysis of the genes involved in eye morphogenesis and development revealed the loss of some conserved noncoding elements (CNEs), which might be associated with the small eyes in catfish. Taken together, our study provided important clues for the adaptation of the catfish visual system to the nocturnal and benthic lifestyles. The draft genome of S. meridionalis represents a valuable resource for studies of the molecular mechanisms of ecological adaptation.

2.
Bioessays ; : e2000161, 2020 Dec 07.
Artigo em Inglês | MEDLINE | ID: mdl-33283342

RESUMO

Recent studies have revealed an astonishing diversity of sex chromosomes in many vertebrate lineages, prompting questions about the mechanisms of sex chromosome turnover. While there is considerable population genetic theory about the evolutionary forces promoting sex chromosome replacement, this theory has not yet been integrated with our understanding of the molecular and developmental genetics of sex determination. Here, we review recent data to examine four questions about how the structure of gene networks influences the evolution of sex determination. We argue that patterns of epistasis, arising from the structure of genetic networks, may play an important role in regulating the rates and patterns of sex chromosome replacement.

3.
Mol Biol Evol ; 2020 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-33300980

RESUMO

Chromosome size and morphology vary within and among species, but little is known about the proximate or ultimate causes of these differences. Cichlid fish species in the tribe Oreochromini share an unusual giant chromosome that is ∼3 times longer than the other chromosomes. This giant chromosome functions as a sex chromosome in some of these species. We test two hypotheses of how this giant sex chromosome may have evolved. The first hypothesis proposes that it evolved by accumulating repetitive elements as recombination was reduced around a dominant sex determination locus, as suggested by canonical models of sex chromosome evolution. An alternative hypothesis is that the giant sex chromosome originated via the fusion of an autosome with a highly repetitive B chromosome, one of which carried a sex determination locus. We test these hypotheses using comparative analysis of chromosome-scale cichlid and teleost genomes. We find that the giant sex chromosome consists of three distinct regions based on patterns of recombination, gene and transposable element content, and synteny to the ancestral autosome. The WZ sex determination locus encompasses the last ∼105Mbp of the 134Mbp giant chromosome. The last 47Mbp of the giant chromosome shares no obvious homology to any ancestral chromosome. Comparisons across 69 teleost genomes reveal that the giant sex chromosome contains unparalleled amounts of endogenous retroviral elements, immunoglobulin genes, and long non-coding RNAs. The results favor the B chromosome fusion hypothesis for the origin of the giant chromosome.

4.
J Periodontol ; 2020 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-33315240

RESUMO

BACKGROUND: Increasing age is associated with systemic diseases as well as with periodontal diseases. We wondered whether a biological age score constructed exclusively from systemic biomarkers would reflect periodontal risk factors at baseline and tooth loss as well as periodontal outcome during 10 years follow-up. METHODS: From the Study of Health in Pomerania (SHIP) 2256 participants (1072 male, 1184 female) were studied for the relationship of the systemic biomarkers glycated hemoglobin (HbA1c), low density lipoprotein cholesterol (LDL), fibrinogen, white blood cell count, blood pressure, and waist circumference to their age. Construction of a biological age (BA) score allowed its comparison with the participants' actual chronological age (CA) and their predisposition to periodontal disease. RESULTS: Though nearly identical in CA, participants appearing younger than their true age had a significantly reduced burden of periodontal risk factors. If BA > CA, then risk factors were more frequent including smoking, oral hygiene, dental visits, education, and income. After 10 years, in participants with identical CA, tooth loss followed their BA calculated at baseline, that is, with BA > CA fewer teeth were preserved. Similarly, periodontal measures varied according to BA; sex differences were obvious. Most significant were BA-related differences in inflammatory and anthropometry parameters. CONCLUSIONS: The results support the assumption that risk profiles aggregated in BA constitute a characteristic susceptibility pattern unique to each individual, common to both systemic and periodontal diseases. Although BA was constructed exclusively from systemic measures at baseline, BA reflects the oral conditions at follow-up.

5.
J Orofac Orthop ; 2020 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-33337523

RESUMO

PURPOSE: Only a few but conflicting results have been reported on the association between malocclusions and caries. We investigated this association using data from the population-based cross-sectional Study of Health in Pomerania (SHIP). METHODS: Sagittal, vertical and transversal intermaxillary relationship, space conditions and sociodemographic parameters of 1210 dentate subjects (median age 30 years, interquartile range 25-35 years) were collected. Caries was assessed with the Decayed-Missing-Filled Surfaces index but analyzed as ordered outcome (four levels: sound, enamel caries, caries, tooth loss) in ordinal multilevel models, taking into account subject, jaw, and tooth level simultaneously. RESULTS: Anterior open bite ≤3 mm (odds ratio [OR] = 2.08, 95% confidence interval [CI]: 1.19-3.61), increased sagittal overjet of 4-6 mm (OR = 1.31, CI: 1.05-1.64), distal occlusion of ½ premolar width (OR = 1.27, CI: 1.05-1.53) and distal 1 premolar width (OR = 1.31, CI: 1.06-1.63) were associated with adjusted increased odds for a higher outcome level (caries). Anterior spacing (OR = 0.24, CI: 0.17-0.33), posterior spacing, (OR = 0.69, CI: 0.5-0.95), posterior crowding (OR = 0.57, CI: 0.49-0.66) and buccal nonocclusion (OR = 0.54, CI: 0.33-0.87) were associated with a lower outcome level (caries). CONCLUSION: The results from this population-based study suggest that a connection between caries and malocclusion exists to a limited extent in young adults. The associations with caries are contradictory for several malocclusion variables. Distal occlusion (OR = 1.31, CI: 1.06-1.63) and related skeletal anomalies displayed positive associations with caries whereas crowding did not. Orthodontic treatment of anterior crowding would probably not interfere with caries experience. These aspects should be considered for patient information and in treatment decisions.

6.
Immunity ; 2020 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-33058782

RESUMO

Several classes of antibiotics have long been known to have beneficial effects that cannot be explained strictly on the basis of their capacity to control the infectious agent. Here, we report that tetracycline antibiotics, which target the mitoribosome, protected against sepsis without affecting the pathogen load. Mechanistically, we found that mitochondrial inhibition of protein synthesis perturbed the electron transport chain (ETC) decreasing tissue damage in the lung and increasing fatty acid oxidation and glucocorticoid sensitivity in the liver. Using a liver-specific partial and acute deletion of Crif1, a critical mitoribosomal component for protein synthesis, we found that mice were protected against sepsis, an observation that was phenocopied by the transient inhibition of complex I of the ETC by phenformin. Together, we demonstrate that mitoribosome-targeting antibiotics are beneficial beyond their antibacterial activity and that mitochondrial protein synthesis inhibition leading to ETC perturbation is a mechanism for the induction of disease tolerance.

7.
Mol Ecol ; 29(24): 4956-4969, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33049090

RESUMO

African cichlid fishes are a prime model for studying speciation mechanisms. Despite the development of extensive genomic resources, it has been difficult to determine which sources of genetic variation are responsible for cichlid phenotypic variation. One of their most variable phenotypes is visual sensitivity, with some of the largest spectral shifts among vertebrates. These shifts arise primarily from differential expression of seven cone opsin genes. By mapping expression quantitative trait loci (eQTL) in intergeneric crosses of Lake Malawi cichlids, we previously identified four causative genetic variants that correspond to indels in the promoters of either key transcription factors or an opsin gene. In this comprehensive study, we show that these indels are the result of the movement of transposable elements (TEs) that correlate with opsin expression variation across the Malawi flock. In tracking the evolutionary history of these particular indels, we found they are endemic to Lake Malawi, suggesting that these TEs are recently active and are segregating within the Malawi cichlid lineage. However, an independent indel has arisen at a similar genomic location in one locus outside of the Malawi flock. The convergence in TE movement suggests these loci are primed for TE insertion and subsequent deletions. Increased TE mobility may be associated with interspecific hybridization, which disrupts mechanisms of TE suppression. This might provide a link between cichlid hybridization and accelerated regulatory variation. Overall, our study suggests that TEs may be an important driver of key regulatory changes, facilitating rapid phenotypic change and possibly speciation in African cichlids.

8.
Genes (Basel) ; 11(9)2020 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-32872430

RESUMO

In domesticated strains of the Nile tilapia, phenotypic sex has been linked to genetic variants on linkage groups 1, 20 and 23. This diversity of sex-loci might reflect a naturally polymorphic sex determination system in Nile tilapia, or it might be an artefact arising from the process of domestication. Here, we searched for sex-determiners in wild populations from Kpandu, Lake Volta (Ghana-West Africa), and from Lake Koka (Ethiopia-East Africa) that have not been subjected to any genetic manipulation. We analysed lab-reared families using double-digest Restriction Associated DNA sequencing (ddRAD) and analysed wild-caught males and females with pooled whole-genome sequencing (WGS). Strong sex-linked signals were found on LG23 in both populations, and sex-linked signals with LG3 were observed in Kpandu samples. WGS uncovered blocks of high sequence coverage, suggesting the presence of B chromosomes. We confirmed the existence of a tandem amh duplication in LG23 in both populations and determined its breakpoints between the oaz1 and dot1l genes. We found two common deletions of ~5 kb in males and confirmed the presence of both amhY and amh∆Y genes. Males from Lake Koka lack both the previously reported 234 bp deletion and the 5 bp frameshift-insertion that creates a premature stop codon in amh∆Y.

9.
Clin Oral Investig ; 2020 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-32964310

RESUMO

OBJECTIVE: SLC22A4/5 single nucleotide polymorphisms (SNPs) have been reported to affect inflammatory diseases. We report the relationship of these polymorphisms with adiposity and tooth loss as elucidated in a 10-year follow-up study. METHODS: Participants of the Study of Health in Pomerania (SHIP, N = 4105) were genotyped for the polymorphisms c.1507C > T in SLC22A4 (rs1050152) and -207C > G in SLC22A5 (rs2631367) using allele-specific real-time PCR assays. A total of 1817 subjects, 934 female and 883 male aged 30-80 years, underwent follow-up 10 years later (SHIP-2) and were assessed for adiposity and tooth loss. RESULTS: The frequencies of the rarer SLC22A4 TT and SLC22A5 CC alleles were 16.7% and 20.3%, respectively. In women, tooth loss was associated with genotype TT vs. CC with incidence rate ratio IRR = 0.74 (95%C.I. 0.60-0.92) and CC vs. GG IRR = 0.79 (0.65-0.96) for SLC22A4 and SLC22A5 SNPs, respectively. In men, no such associations were observed. In the follow-up examination, the relationship between tooth loss and these SNPs was in parallel with measures of body shape such as BMI, body weight, waist circumference, or body fat accumulation. The association between muscle strength and body fat mass was modified by the genotypes studied. CONCLUSIONS: SLC22A4 c.150C > T and SLC22A5 -207C > G polymorphisms are associated with tooth loss and markers of body shape in women but not in men. CLINICAL RELEVANCE: Tooth loss may be related to obesity beyond inflammatory mechanisms, conceivably with a genetic background.

10.
Cell ; 182(6): 1490-1507.e19, 2020 Sep 17.
Artigo em Inglês | MEDLINE | ID: mdl-32916131

RESUMO

Metabolic reprogramming is a key feature of many cancers, but how and when it contributes to tumorigenesis remains unclear. Here we demonstrate that metabolic reprogramming induced by mitochondrial fusion can be rate-limiting for immortalization of tumor-initiating cells (TICs) and trigger their irreversible dedication to tumorigenesis. Using single-cell transcriptomics, we find that Drosophila brain tumors contain a rapidly dividing stem cell population defined by upregulation of oxidative phosphorylation (OxPhos). We combine targeted metabolomics and in vivo genetic screening to demonstrate that OxPhos is required for tumor cell immortalization but dispensable in neural stem cells (NSCs) giving rise to tumors. Employing an in vivo NADH/NAD+ sensor, we show that NSCs precisely increase OxPhos during immortalization. Blocking OxPhos or mitochondrial fusion stalls TICs in quiescence and prevents tumorigenesis through impaired NAD+ regeneration. Our work establishes a unique connection between cellular metabolism and immortalization of tumor-initiating cells.

11.
J Biophotonics ; : e202000221, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32931142

RESUMO

Reinfection in endodontically treated teeth is linked to the complexity of the root canal system, which is problematic to reach with conventional disinfection methods. As plasma is expected to have the ability to sanitize narrow areas, the aim of this study was to analyze the effect of cold atmospheric pressure plasma (CAP) on Candida albicans in root canals of extracted human teeth. CAP was applied as mono treatment and in combination with standard endodontic disinfectants (sodium hypochlorite, chlorhexidine and octenidine). Disinfection efficiency was evaluated as reduction of the logarithm of colony forming units per milliliter (log10 CFU/mL) supported by scanning electron microscopy as imaging technique. Plasma alone showed the highest reduction of log10 CFU, suggesting the best disinfection properties of all tested agents.

12.
BMC Evol Biol ; 20(1): 95, 2020 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-32736512

RESUMO

BACKGROUND: Adaptive radiations are characterized by extreme and/or iterative phenotypic divergence; however, such variation does not accumulate evenly across an organism. Instead, it is often partitioned into sub-units, or modules, which can differentially respond to selection. While it is recognized that changing the pattern of modularity or the strength of covariation (integration) can influence the range or rate of morphological evolution, the relationship between shape variation and covariation remains unclear. For example, it is possible that rapid phenotypic change requires concomitant changes to the underlying covariance structure. Alternatively, repeated shifts between phenotypic states may be facilitated by a conserved covariance structure. Distinguishing between these scenarios will contribute to a better understanding of the factors that shape biodiversity. Here, we explore these questions using a diverse Lake Malawi cichlid species complex, Tropheops, that appears to partition habitat by depth. RESULTS: We construct a phylogeny of Tropheops populations and use 3D geometric morphometrics to assess the shape of four bones involved in feeding (mandible, pharyngeal jaw, maxilla, pre-maxilla) in populations that inhabit deep versus shallow habitats. We next test numerous modularity hypotheses to understand whether fish at different depths are characterized by conserved or divergent patterns of modularity. We further examine rates of morphological evolution and disparity between habitats and among modules. Finally, we raise a single Tropheops species in environments mimicking deep or shallow habitats to discover whether plasticity can replicate the pattern of morphology, disparity, or modularity observed in natural populations. CONCLUSIONS: Our data support the hypothesis that conserved patterns of modularity permit the evolution of divergent morphologies and may facilitate the repeated transitions between habitats. In addition, we find the lab-reared populations replicate many trends in the natural populations, which suggests that plasticity may be an important force in initiating depth transitions, priming the feeding apparatus for evolutionary change.


Assuntos
Ciclídeos/anatomia & histologia , Ecossistema , Comportamento Alimentar , Animais , Arcada Osseodentária/anatomia & histologia , Lagos , Malaui , Mandíbula/anatomia & histologia , Modelos Biológicos , Faringe/anatomia & histologia , Filogenia , Água
13.
Clin Oral Investig ; 2020 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-32827080

RESUMO

OBJECTIVES: We aimed at investigating whether the interaction between the local inflammation, periodontitis, and obesity is independently associated with systemic inflammation. METHODS: From the population-based Study of Health in Pomerania, 3366 participants, without (2366) and with (1000) obesity, were studied for the association of periodontitis, measured as probing depth (PD) and plaque together with body mass index (BMI) on C-reactive protein (CRP). Quantile regression was used to evaluate the association between periodontal, anthropometric, and inflammatory variables (outcomes). RESULTS: The overall prevalence of obesity in this adult population was 31.4% in men and 28.1% in women. Both PD and plaque were positively associated with CRP, revealing an increasing impact across the CRP concentration distribution. Adjusting the regression of CRP or fibrinogen on PD for waist circumference attenuated but did not abolish the PD coefficients. Dental plaque was similarly associated with these interrelations. Association between PD and a dental plaque was different among participants with low-, medium-, or high-risk CRP concentrations. CONCLUSION: Local and systemic sources of inflammation contribute to blood levels of inflammatory markers. The respective contributions depend on the relative rate in each of the inflammation-inducing risks and are dominated by adiposity. CLINICAL RELEVANCE: Keeping systemic inflammation low in order to prevent age-related disease sequelae.

14.
Artigo em Inglês | MEDLINE | ID: mdl-32477961

RESUMO

The aim of this follow-up study was, to compare the effects of mechanical periodontal therapy with or without adjunctive amoxicillin and metronidazole on the subgingival microbiome of smokers with periodontitis using 16S rDNA amplicon next generation sequencing. Fifty-four periodontitis patients that smoke received either non-surgical periodontal therapy with adjunctive amoxicillin and metronidazole (n = 27) or with placebos (n = 27). Subgingival plaque samples were taken before and two months after therapy. Bacterial genomic DNA was isolated and the V4 hypervariable region of the bacterial 16S rRNA genes was amplified. Up to 96 libraries were normalized and pooled for Illumina MiSeq paired-end sequencing with almost fully overlapping 250 base pairs reads. Exact ribosomal sequence variants (RSVs) were inferred with DADA2. Microbial diversity and changes on the genus and RSV level were analyzed with non-parametric tests and a negative binomial regression model, respectively. Before therapy, the demographic, clinical, and microbial parameters were not significantly different between the placebo and antibiotic groups. Two months after the therapy, clinical parameters improved and there was a significantly increased dissimilarity of microbiomes between the two groups. In the antibiotic group, there was a significant reduction of genera classified as Porphyromonas, Tannerella, and Treponema, and 22 other genera also decreased significantly, while Selenomonas, Capnocytophaga, Actinomycetes, and five other genera significantly increased. In the placebo group, however, there was not a significant decrease in periodontal pathogens after therapy and only five other genera decreased, while Veillonella and nine other genera increased. We conclude that in periodontitis patients who smoke, microbial shifts occurred two months after periodontal therapy with either antibiotics or placebo, but genera including periodontal pathogens decreased significantly only with adjunctive antibiotics.

15.
Mol Ecol Resour ; 20(5): 1361-1371, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32419357

RESUMO

Onychostoma macrolepis is an emerging commercial cyprinid fish species. It is a model system for studies of sexual dimorphism and genome evolution. Here, we report the chromosome-level assembly of the O.macrolepis genome obtained from the integration of nanopore long-read sequencing with physical maps produced using Bionano and Hi-C technology. A total of 87.9 Gb of nanopore sequence provided approximately 100-fold coverage of the genome. The preliminary genome assembly was 883.2 Mb in size with a contig N50 size of 11.2 Mb. The 969 corrected contigs obtained from Bionano optical mapping were assembled into 853 scaffolds and produced an assembly of 886.5 Mb with a scaffold N50 of 16.5 Mb. Finally, using the Hi-C data, 881.3 Mb (99.4% of genome) in 526 scaffolds were anchored and oriented in 25 chromosomes ranging in size from 25.27 to 56.49 Mb. In total, 24,770 protein-coding genes were predicted in the genome, and ~96.85% of the genes were functionally annotated. The annotated assembly contains 93.3% complete genes from the BUSCO reference set. In addition, we identified 409 Mb (46.23% of the genome) of repetitive sequence, and 11,213 non-coding RNAs, in the genome. Evolutionary analysis revealed that O. macrolepis diverged from common carp approximately 24.25 million years ago. The chromosomes of O. macrolepis showed an unambiguous correspondence to the chromosomes of zebrafish. The high-quality genome assembled in this work provides a valuable genomic resource for further biological and evolutionary studies of O. macrolepis.

16.
J Int Acad Periodontol ; 22(2): 41-53, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32224549

RESUMO

OBJECTIVE: The selection of proper outcome measures is a critical step in clinical research. Most randomized clinical trials (RCTs) assessing the effects of initial anti-infective periodontal therapies use surrogate outcomes as primary outcome variables, such as mean changes in probing depth (PD) or in clinical attachment. However, these parameters do not reflect disease remission/control at patient level, which has led to subjective interpretations of the data from RCTs and Systematic Reviews. Based on a comprehensive analysis of 724 patients from USA, Germany and Brazil treated for periodontitis, this paper suggests that the clinical endpoint of "≤4 sites with PD≥5mm" is effective in determining disease remission/control after active periodontal treatment and therefore, may represent a pertinent endpoint for applying the treat-to-target concept in RCTs. Furthermore, regression models showed that the presence of >10% and >20% sites with bleeding on probing in the mouth post-treatment increases the risk of a patient leaving the endpoint from 1-2 years (OR=3.5 and 8.7, respectively). Researchers are encouraged to present results on this outcome when reporting their trials, as this will allow for an objective comparison across studies and facilitate systematic reviews, and consequently, the extrapolation of data from research to clinical practice.


Assuntos
Periodontite , Brasil , Alemanha , Humanos , Ensaios Clínicos Controlados Aleatórios como Assunto , Revisões Sistemáticas como Assunto
17.
J Clin Periodontol ; 47(7): 842-850, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32248552

RESUMO

AIM: The aim of this study was to evaluate the effect of non-surgical periodontal therapy on circulating levels of the systemic inflammation-associated biomarkers orosomucoid (ORM), high-sensitivity C-reactive protein (hsCRP), chemerin, and retinol-binding protein 4 (RBP4) in overweight or normal-weight patients with periodontitis at 27.5 months after therapy. MATERIALS AND METHODS: This exploratory subanalysis includes patients from the ABPARO-trial (ClinicalTrials.gov NCT00707369). The per-protocol collective provided untreated periodontitis patients with high (≥28 kg/m2 ) or moderate (21-24 kg/m2 ) BMI. Out of the per-protocol collective, 80 patients were randomly selected and stratified for BMI group, sex, and treatment group (antibiotics/placebo), resulting in 40 overweight and normal-weight patients. Patients received non-surgical periodontal therapy and maintenance at 3-month intervals. Plasma samples from baseline and 27.5 months following initial treatment were used to measure the concentrations of ORM, hsCRP, chemerin, and RBP4. RESULTS: At the 27.5-month examination, ORM and hsCRP decreased noticeably in the overweight group (ORM: p = .001, hsCRP: p = .004) and normal-weight patients (ORM: p = .007, hsCRP: p < .001). Chemerin decreased in the overweight group (p = .048), and RBP4 concentrations remained stable. CONCLUSION: Non-surgical periodontal therapy reduced systemically elevated inflammation-associated biomarkers in periodontitis patients. These improvements were more pronounced in overweight patients than in normal-weight patients.

18.
Cell Commun Signal ; 18(1): 61, 2020 04 10.
Artigo em Inglês | MEDLINE | ID: mdl-32276641

RESUMO

BACKGROUND: Cutaneous squamous cell carcinomas (cSCC) are the primary cause of premature deaths in patients suffering from the rare skin-fragility disorder recessive dystrophic epidermolysis bullosa (RDEB), which is in marked contrast to the rarely metastasizing nature of these carcinomas in the general population. This remarkable difference is attributed to the frequent development of chronic wounds caused by impaired skin integrity. However, the specific molecular and cellular changes to malignancy, and whether there are common players in different types of aggressive cSCCs, remain relatively undefined. METHODS: MiRNA expression profiling was performed across various cell types isolated from skin and cSCCs. Microarray results were confirmed by qPCR and by an optimized in situ hybridization protocol. Functional impact of overexpression or knock-out of a dysregulated miRNA was assessed in migration and 3D-spheroid assays. Sample-matched transcriptome data was generated to support the identification of disease relevant miRNA targets. RESULTS: Several miRNAs were identified as dysregulated in cSCCs compared to control skin. These included the metastasis-linked miR-10b, which was significantly upregulated in primary cell cultures and in archival biopsies. At the functional level, overexpression of miR-10b conferred the stem cell-characteristic of 3D-spheroid formation capacity to keratinocytes. Analysis of miR-10b downstream effects identified a novel putative target of miR-10b, the actin- and tubulin cytoskeleton-associated protein DIAPH2. CONCLUSION: The discovery that miR-10b mediates an aspect of cancer stemness - that of enhanced tumor cell adhesion, known to facilitate metastatic colonization - provides an important avenue for future development of novel therapies targeting this metastasis-linked miRNA.

19.
J Invest Dermatol ; 140(10): 1985-1993.e5, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32142798

RESUMO

End-joining‒based gene editing is frequently used for efficient reframing and knockout of target genes. However, the associated random, unpredictable, and often heterogeneous repair outcomes limit its applicability for therapeutic approaches. This study revealed more precise and predictable outcomes simply on the basis of the sequence context at the CRISPR/Cas9 target site. The severe dystrophic form of the blistering skin disease epidermolysis bullosa (DEB) represents a suitable model platform to test these recent developments for the disruption and reframing of dominant and recessive alleles, respectively, both frequently seen in DEB. We delivered a CRISPR/Cas9 nuclease as ribonucleoprotein into primary wild-type and recessive DEB keratinocytes to introduce a precise predictable single adenine sense-strand insertion at the target site. We achieved type VII collagen knockout in more than 40% of ribonucleoprotein-treated primary wild-type keratinocytes and type VII collagen restoration in more than 70% of ribonucleoprotein-treated recessive DEB keratinocytes. Next-generation sequencing of the on-target site revealed the presence of the precise adenine insertion upstream of the pathogenic mutation in at least 17% of all analyzed COL7A1 alleles. This demonstrates that COL7A1 editing based on precise end-joining‒mediated DNA repair is an efficient strategy to revert the disease-associated nature of DEB regardless of the mutational inheritance.

20.
Artigo em Alemão | MEDLINE | ID: mdl-32166336

RESUMO

BACKGROUND: Caries and periodontitis are highly prevalent worldwide. Because detailed data on these oral diseases were collected within the framework of the German National Cohort (GNC), associations between oral and systemic diseases and conditions can be investigated. OBJECTIVES: The study protocol for the oral examination was designed to ensure a comprehensive collection of dental findings by trained non-dental staff within a limited examination time. At the mid-term of the GNC baseline examination, a first quality evaluation was performed to check the plausibility of results and to propose measures to improve the data quality. MATERIALS AND METHODS: A dental interview, saliva sampling and oral diagnostics were conducted. As part of the level­1 examination, the number of teeth and prostheses were recorded. As part of the level­2 examination, detailed periodontal, cariological and functional aspects were examined. All examinations were conducted by trained non-dental personnel. Parameters were checked for plausibility and variable distributions were descriptively analysed. RESULTS: Analyses included data of 57,967 interview participants, 56,913 level­1 participants and 6295 level­2 participants. Percentages of missing values for individual clinical parameters assessed in level 1 and level 2 ranged between 0.02 and 3.9%. Results showed a plausible distribution of the data; rarely, implausible values were observed, e.g. for measurements of horizontal and vertical overbite (overjet and overbite). Intra-class correlation coefficients indicated differences in individual parameters between regional clusters, study centres and across different examiners. CONCLUSIONS: The results confirm the feasibility of the study protocol by non-dental personnel and its successful integration into the GNC's overall assessment program. However, rigorous dental support of the study centres is required for quality management.


Assuntos
Coleta de Dados/normas , Cárie Dentária , Doenças da Boca , Saúde Bucal , Estudos de Coortes , Cárie Dentária/epidemiologia , Alemanha , Humanos , Garantia da Qualidade dos Cuidados de Saúde , Controle de Qualidade
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