mtDNA diversity in human populations highlights the merit of haplotype matching in gene therapies

STUDY QUESTION Does mitochondrial DNA (mtDNA) diversity in modern human populations potentially pose a challenge, via mtDNA segregation, to mitochondrial replacement therapies? SUMMARY ANSWER The magnitude of mtDNA diversity in modern human populations is as high as in mammalian model systems where strong mtDNA segregation is observed; consideration of haplotype pairs and/or haplotype matching can help avoid these potentially deleterious effects. WHAT IS KNOWN ALREADY In mammalian models, substantial proliferative differences are observed between different mtDNA haplotypes in cellular admixtures, with larger proliferative differences arising from more diverse haplotype pairings. If maternal mtDNA is ‘carried over’ in human gene therapies, these proliferative differences could lead to its amplification in the resulting offspring, potentially leading to manifestation of the disease that the therapy was designed to avoid—but existing studies have not investigated whether mtDNA diversity in modern human populations is sufficient to permit significant amplification. STUDY DESIGN, SIZE, DURATION This theoretical study used over 7500 human mtDNA sequences from The National Center for Biotechnology Information (NCBI), a range of international and British mtDNA surveys, and 2011 census data. PARTICIPANTS/MATERIALS, SETTING, METHODS A stochastic simulation approach was used to model random haplotype pairings from within different regions. In total, 1000 simulated pairings were analysed using the basic local alignment search tool (BLAST) for each region. Previous data from mouse models were used to estimate proliferative differences. MAIN RESULTS AND THE ROLE OF CHANCE Even within the same haplogroup, differences of around 20–80 single-nucleotide polymorphisms (SNPs) are common between mtDNAs admixed in random pairings. These values are sufficient to lead to substantial segregation in mouse models over an organismal lifetime, even given low starting heteroplasmy, inducing increases from 5% to 35% over 1 year. Substantial population mixing in modern UK cities increases the expected genetic differences. Hence, the likely genetic differences between humans randomly sampled from a population may well allow substantial amplification of a disease-carrying mtDNA haplotype over the timescale of a human lifetime. We report ranges and mean differences for all statistics to quantify uncertainty in our results. LIMITATIONS/REASONS FOR CAUTION The mapping from mouse and other mammalian models to the human system is challenging, as timescales and mechanisms may differ. Reporting biases in NCBI mtDNA data, if present, may affect the statistics we compute. We discuss the robustness of our findings in the light of these concerns. WIDER IMPLICATIONS OF THE FINDINGS Matching the mtDNA haplotypes of the mother and third-party donor in mitochondrial replacement therapies is supported as a means of ameliorating the potentially deleterious results of human mtDNA diversity. We present a chart of expected SNP differences between mtDNA haplogroups, allowing the selection of optimal partners for therapies

Time- and compartment-resolved proteome profiling of the extracellular niche in lung injury and repair

The extracellular matrix (ECM) is a key regulator of tissue morphogenesis and repair. However, its composition and architecture are not well characterized. Here, we monitor remodeling of the extracellular niche in tissue repair in the bleomycin-induced lung injury mouse model. Mass spectrometry quantified 8,366 proteins from total tissue and bronchoalveolar lavage fluid (BALF) over the course of 8 weeks, surveying tissue composition from the onset of inflammation and fibrosis to its full recovery. Combined analysis ofproteome, secretome, and transcriptome highlighted post-transcriptional events during tissue fibrogenesis and defined the composition of airway epithelial lining fluid. To comprehensively characterize the ECM, we developed a quantitative detergent solubility profiling (QDSP) method, which identified Emilin-2 and collagen-XXVIII as novel constituents of the provisional repair matrix. QDSP revealed which secreted proteins interact with the ECM, and showed drastically altered association of morphogens to the insoluble matrix upon injury. Thus, our proteomic systems biology study assigns proteins to tissue compartments and uncovers their dynamic regulation upon lung injury and repair, potentially contributing to the development of anti-fibrotic strategies

Hierarchical carbon fibre composites incorporating high loadings of carbon nanotubes

Uncured solid bisphenol-A epoxy resins containing up to 20 wt% carbon nanotubes (CNTs) were prepared using melt blending in a high shear mixer. The extrudate was ground to produce fine nanocomposite (NC) powders. This simple method produced well-dispersed NC, with CNT agglomerate sizes below 1 μm. Consolidated NCs displayed improved tensile moduli and strengths up to 3.3 GPa (+32%) and 78 MPa (+19%), respectively at 15 wt% CNT, compared to the pure cured epoxy matrix. The relatively high Tg of 39 ◦C for the uncured NC powders simplified the manufacture of composite prepregs using wet powder impregnation. The prepregs were laminated into hierarchical carbon fibre reinforced composites with improved through-thickness properties. Interlaminar shear strength improved for intermediate CNT loadings in the matrix up to 65 MPa (10 wt% CNT, +19%) but decreased at higher concentrations. Compression moduli remained constant irrespectively of CNT loading but compression strength increased with a CNT loading of 2.5 wt% to 772 MPa (+31%). The mechanical properties of the hierarchical composites reflect good consolidation (void content <3%) and excellent fibre alignment (<±0.8◦). In addition to the improved mechanical properties, incorporation of CNTs improved the through- thickness electrical conductivity up to 115 S/

Simulating the recycling of milk bottles in the UK: Influence of blending virgin and repeatedly melt-extruded high-density polyethylene

The UK Dairy Roadmap has set a target of achieving 50 wt.-% high density polyethylene (HDPE) recyclate in their HDPE milk bottles. Such high recyclate content will lead to the accumulation of HDPE recyclates that have been subjected to different number of melt extrusion cycles in the supply chain. This work investigates the structure-property relationship of blending virgin HDPE (vHDPE) with these different grades of repeatedly melt-extruded HDPE (rHDPE). HDPE was subjected to 10, 20 and 50 melt-extrusion cycles and blended with vHDPE. No significant difference in terms of melt rheology, tensile properties and overall migration in acidic and aqueous environments of the blends of the different rHDPEs with vHDPE was observed when compared to vHDPE. This study demonstrates the feasibility of blending up to 50 wt.-% rHDPE of different grades with vHDPE as set out in the UK Dairy Roadmap

Agonist-mediated switching of ion selectivity in TPC2 differentially promotes lysosomal function

Ion selectivity is a defining feature of a given ion channel and is considered immutable. Here we show that ion selectivity of the lysosomal ion channel TPC2, which is hotly debated (Calcraft et al., 2009; Guo et al., 2017; Jha et al., 2014; Ruas et al., 2015; Wang et al., 2012), depends on the activating ligand. A high-throughput screen identified two structurally distinct TPC2 agonists. One of these evoked robust Ca2+-signals and non-selective cation currents, the other weaker Ca2+-signals and Na+-selective currents. These properties were mirrored by the Ca2+-mobilizing messenger, NAADP and the phosphoinositide, PI(3,5)P2, respectively. Agonist action was differentially inhibited by mutation of a single TPC2 residue and coupled to opposing changes in lysosomal pH and exocytosis. Our findings resolve conflicting reports on the permeability and gating properties of TPC2 and they establish a new paradigm whereby a single ion channel mediates distinct, functionally-relevant ionic signatures on demand

Lasp-1 Regulates Podosome Function

Eukaryotic cells form a variety of adhesive structures to connect with their environment and to regulate cell motility. In contrast to classical focal adhesions, podosomes, highly dynamic structures of different cell types, are actively engaged in matrix remodelling and degradation. Podosomes are composed of an actin-rich core region surrounded by a ring-like structure containing signalling molecules, motor proteins as well as cytoskeleton-associated proteins

Health-related quality of life in lower-risk MDS patients compared with age- and sex-matched reference populations: a European LeukemiaNet study

In myelodysplastic syndromes (MDS), health-related quality of life (HRQoL) represents a relevant patient-reported outcome, which is essential in individualized therapy planning. Prospective data on HRQoL in lower-risk MDS remain rare. We assessed HRQOL by EQ-5D questionnaire at initial diagnosis in 1690 consecutive IPSS-Low/Int-1 MDS patients from the European LeukemiaNet Registry. Impairments were compared with age- and sex-matched EuroQol Group norms. A significant proportion of MDS patients reported moderate/severe problems in the dimensions pain/discomfort (49.5%), mobility (41.0%), anxiety/depression (37.9%), and usual activities (36.1%). Limitations in mobility, self-care, usual activities, pain/discomfort, and EQ-VAS were significantly more frequent in the old, in females, and in those with high co-morbidity burden, low haemoglobin levels, or red blood cells transfusion need (

Expression of RHOGTPase regulators in human myometrium

<p>Abstract</p> <p>Background</p> <p>RHOGTPases play a significant role in modulating myometrial contractility in uterine smooth muscle. They are regulated by at least three families of proteins, RHO guanine nucleotide exchange factors (RHOGEFs), RHOGTPase-activating proteins (RHOGAPs) and RHO guanine nucleotide inhibitors (RHOGDIs). RHOGEFs activate RHOGTPases from the inactive GDP-bound to the active GTP-bound form. RHOGAPs deactivate RHOGTPases by accelerating the intrinsic GTPase activity of the RHOGTPases, converting them from the active to the inactive form. RHOGDIs bind to GDP-bound RHOGTPases and sequester them in the cytosol, thereby inhibiting their activity. Ezrin-Radixin-Moesin (ERM) proteins regulate the cortical actin cytoskeleton, and an ERM protein, moesin (MSN), is activated by and can also activate RHOGTPases.</p> <p>Methods</p> <p>We therefore investigated the expression of various RHOGEFs, RHOGAPs, a RHOGDI and MSN in human myometrium, by semi-quantitative reverse transcription PCR, real-time fluorescence RT-PCR, western blotting and immunofluorescence microscopy. Expression of these molecules was also examined in myometrial smooth muscle cells.</p> <p>Results</p> <p>ARHGEF1, ARHGEF11, ARHGEF12, ARHGAP5, ARHGAP24, ARHGDIA and MSN mRNA and protein expression was confirmed in human myometrium at term pregnancy, at labour and in the non-pregnant state. Furthermore, their expression was detected in myometrial smooth muscle cells. It was determined that ARHGAP24 mRNA expression significantly increased at labour in comparison to the non-labour state.</p> <p>Conclusion</p> <p>This study demonstrated for the first time the expression of the RHOGTPase regulators ARHGEF1, ARHGEF11, ARHGEF12, ARHGAP5, ARHGAP24, ARHGDIA and MSN in human myometrium, at term pregnancy, at labour, in the non-pregnant state and also in myometrial smooth muscle cells. ARHGAP24 mRNA expression significantly increased at labour in comparison to the non-labouring state. Further investigation of these molecules may enable us to further our knowledge of RHOGTPase regulation in human myometrium during pregnancy and labour.</p

Keeping the Vimentin Network under Control: Cell–Matrix Adhesion–associated Plectin 1f Affects Cell Shape and Polarity of Fibroblasts

Mature focal adhesions and fibrillar adhesions act as anchorage sites for vimentin filaments, with plectin isoform 1f being the crucial linker protein. Plectin serves as a nucleation and assembly center for the de novo formation of vimentin networks. Anchored vimentin creates a resilient cage-like core structure that affects cell shape