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Induction of ultra-morphological features of apoptosis in mature and immature sperm

Asian Journal of Andrology 2017年第5期19卷 533-537页

作者： sonja grunewald Guenther Fitzl Christopher Springsguth Department of Dermatology Venerology and Allergology University Hospital Leipzig Leipzig Germany Institute of Anatomy University of Leipzig Leipzig Germany

There is a fundamental body of evidence suggesting that activated apoptosis signaling in ejaculated human sperm negatively influences their fertilization potential. However, it is still controversial whether this apoptotic signaling is a relic of an abortive apoptosis related to spermatogenesis or if it should be regarded as a functional preformed pathway in mature sperm leading to stereotypical morphological changes reflecting nuclear disassembly. To address this question, apoptosis was induced using betulinic acid in mature and immature ejaculated human sperm enriched by density gradient centrifugation. Execution of apoptosis was monitored by observing ultra-morphological changes via transmission electron microscopy. Typical morphological signs of apoptosis in somatic cells include plasma membrane blebbing with the formation of apoptotic bodies, impaired mitochondrial integrity, defects of the nuclear envelope, and nuclear fragmentation; these morphologies have also been observed in human sperm. In addition, these apoptotic characteristics were more frequent in immature sperm compared to mature sperm. Following betulinic acid treatment, apoptosis-related morphological changes were induced in mature sperm from healthy donors. This effect was much less pronounced in immature sperm. Moreover, in both fractions, the betulinic acid treatment increased the percentage of acrosome-reacted sperm. The results of our ultra-morphological study prove the functional competence of apoptosis in mature ejaculated human sperm. The theory of a sole abortive process may be valid only for immature sperm. The induction of the acrosome reaction by stimulating apoptosis might shed light on the biological relevance of sperm apoptosis.

关键词： apoptosis electron microscopy human morphology sperm

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Defective claudin-10 causes a novel variation of HELIX syndrome through compromised tight junction strand assembly

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Genes & Diseases 2022年第5期9卷 1301-1314页

作者： Sebastian Sewerin Jorg Piontek Ria Schonauer sonja grunewald Angelika Rauch Steffen Neuber Carsten Bergmann Dorothee Gunzel Jan Halbritte Division of Nephrology University of Leipzig Medical CenterLeipzig 04103Germany Clinical Physiology/Nutritional Medicine Charitee Universitatsmedizin BerlinBerlin 12203Germany Division of Dermatology Venereologyand AllergologyUniversity of Leipzig Medical CenterLeipzig 04103Germany Division of Prosthodontics and Materials Science University of Leipzig Medical CenterLeipzig 04103Germany Center for Human Genetics BioscientiaIngelheim 55218Germany f Medizinische Genetik MainzLimbach GeneticsMainz 55128Germany

Formation of claudin-10 based tight junctions(TJs)is paramount to paracellular Na+transport in multiple *** variants in CLDN10 have been linked to HELIX syndrome,a salt-losing tubulopathy with altered handling of divalent cations accompanied by dysfunctional salivary,sweat,and lacrimal ***,we investigate molecular basis and phenotypic consequences of a newly identified homozygous CLDN10 variant that translates into a single amino acid substitution within the fourth transmembrane helix of *** addition to hypohidrosis(H),electrolyte(E)imbalance with impaired urine concentrating ability,and hypolacrimia(L),phenotypic findings include altered salivary electrolyte composition and amelogenesis imperfecta but neither ichthyosis(I)nor xerostomia(X).Employing cellular TJ reconstitution assays,we demonstrate perturbation of cis-and trans-interactions between mutant claudin-10 *** of reconstituted TJ strands show disturbed continuity and reduced abundance in the mutant ***,both major isoforms,claudin-10a and claudin-10b,are differentially affected with claudin-10b showing more severe molecular ***,expression of the mutant in renal epithelial cells with endogenous TJs results in wild-type-like ion selectivity and conductivity,indicating that aberrant claudin-10 is generally capable of forming functional paracellular ***,mutant proteins prove pathogenic by compromising claudin-10 TJ strand *** ex vivo investigations indicate their insertion into TJs to occur in a tissue-specific manner.

关键词： Claudin-10 HELIX syndrome Paracellular transport Salt-losing tubulopathy Tight junction

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Immunomagnetic removal of cryo-damaged human spermatozoa

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Asian Journal of Andrology 2005年第1期7卷 61-69,页

作者： Uwe Paasch sonja grunewald Katja Wuendrich Tbrsten Jope Hans-Jurgen Glander Division of Andrology Department of Dermatology University of Leipzig Leipzig D-04103 Germany

Aim: To estimate the dissipation of mitochondrial transmembrane potential (mTMR,Δψ_m) and activation of sperm caspases (aCP) as signs of apoptosis in human spermatozoa during cryopreservation and to evaluate the efficiency of immunomagnetic cell separation (MACS) of these spermatozoa via annexin V-binding. Methods: The mTMP and aCP in fresh and cryopreserved spermatozoa were detected by fluorescence microscopy and by Western blots. The sperm suspensions were divided into two sperm fractions (with intact and deteriorated membranes) by magnetic cell separation (MiniMACS, Miltenyi Biotec, Bergisch Gladbach, Germany) in dependence on their binding to superparamagnetic annexin V-microbeads (AN-MB). Results: The cryopreservation decreased the portion of spermatozoa with intact mTMP from 80.1% ± 7.2 % to 53.5 % ± 13.1% and increased the spermatozoa with activated pancaspases (aCP) from 21.8 % ± 2.6 % to 47.7 % ± 5.8 % (n = 10; mean ± SEM; P < 0.01). The activation of caspases 1, 3, 8, and 9 in the cryopreserved spermatozoa was confirmed by Western blots (n = 22). MACS reduced significantly the percentage of cryopreserved spermatozoa with dissipated mTMP to 8.1 ± 3.9 (P < 0.01) and also those with aCP to 9.3 % ± 2.2 %. Western blot analyses confirmed the increase of the activated caspase3, 9, and 8 in the AN-MB-positive fraction (P < 0.05) compared with the AN-MB-negative fraction. The MACS separation effect was confirmed by anti-annexin V-antibodies. There was no significant influence of the separation column and the magnetic field on the sperm functions. Conclusion: The cryopreservation impaired the mTMP and enhanced the activation status of caspases in human spermatozoa. The immunomagnetic sperm separation via binding of AN-MB could deplete low quality spermatozoa from cryopreserved semen samples.

关键词： spermatozoa cryopreservation immunomagnetic separation plasma membrane annexin V apoptosis caspases mitochondria

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Stability of fluorochrome based assays to measure subcellular sperm functions

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Asian Journal of Andrology 2008年第3期10卷 455-459页

作者： sonja grunewald Manja Rasch Martin Reinhardt Thomas Baumann Uwe Paasch Hans-Juergen Glander Department of Dermatology/Andrology Unit European Academy of Andrology （EAA ） Training Center University of Leipzig Leipzig 04103 Germany

Aim： To evaluate the long-term stability of the fluorescence signals of new fluorescence-based semen analysis assays for clinical application. Methods： Semen samples from 87 unselected infertile patients were used to perform the following assays：（i） detection of active caspase-3 （n = 17）; （ii） integrity of the mitochondrial membrane potential （MMP）（n = 17）; （iii） externalization of phosphatidylserine （EPS）（n = 16）; and （iv） detection of intact acrosomes via CD46 （n = 37）. After the assays, 4% paraformaldehyde was added to all aliquots. The fluorescence intensity of each sample was evaluated by flow cytometry on days 0, 3, 7, 10 and 14. Results： Differences of up to ± 5% positive spermatozoa from the value measured at day 0 were estimated as acceptable deviation. The Caspase-3 FLICA^TM showed mean differences 〈 5% at day 3, 7 and 10. At day 14 the mean difference was 7.6%. In contrast, the disrupted MMP and the EPS detection showed differences 〉 5% at day 3. The CD46-FITC labeling displayed absolute differences 〈 5% CD46-positive spermatozoa at days 3, 7, 10 and 14. Conclusion： Although immediate analysis of the fluorescence signals is recommended, it is possible to evaluate caspase-3 activation up to 10 days and CD46 up to 14 days after staining of sperm. The FACS evaluation of MMP and EPS detection should be conducted on the same day.

关键词： human spermatozoa apoptosis assays caspase-3 mitochondrial membrane potential integrity CD46 semen analysis

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Cryptic exon inclusion in TDP-43 proteinopathies:opportunities and challenges

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Neural Regeneration Research 2025年第7期20卷 2003-2004页

作者： Lorena Decker sonja Menge Axel Freischmidt Department of Neurology Ulm UniversityUlmGermany

TDP-43 proteinopathies and cryptic exons:Transactive response DNA binding protein of 43 kDa(TDP-43)is a ubiquitously expressed RNA/DNA binding protein crucial for coding and non-coding RNA metabolism including transcription,splicing,transport,translation,and ***-43 shuttles between the nucleus and cytoplasm,but is predominantly localized in the *** diseases(NDs)may be accompanied by nuclear loss and possible cytoplasmic accumulation and aggregation of TDP-43 in vulnerable neurons and *** neuropathology is the hallmark of most individuals suffering from amyotrophic lateral sclerosis(ALS),frontotemporal dementia(FTD)with TDP-43-immunoreactive pathology(FTD-TDP),limbic-predominant age-related TDP-43 encephalopathy(LATE)and Perry syndrome,but also coexists with the primary pathology in subsets of patients suffering from other NDs,such as Alzheimer’s disease,Lewy body dementias,or Huntington’s *** in the gene encoding TDP-43(TARDBP)are the cause of ALS and/or FTD in some rare cases substantiating the importance of this protein in aging *** is still controversial if loss of nuclear,or increased cytoplasmic and/or aggregating TDP-43 is more harmful to neurons(Nag and Schneider,2023).Recently,the role of nuclear TDP-43 in repressing the inclusion of intronic sequences,named cryptic exons(CEs),into mature mRNAs gained much attention.

关键词： Huntington metabolism TDP

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A probabilistic-based method to evaluate hygrothermal performance of an internally insulated brick wall

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Building Simulation 2021年第2期14卷 283-299页

作者： Jianhua Zhao Jianshun Zhang John grunewald Shuo Feng Institute of Architectural Technology&Science School of ArchitectureTianjin UniversityTianjinChina College of Mechanical&Aerospace Engineering Syracuse UniversitySyracuseUSA Institute of Building Climatology School of ArchitectureTechnische Universität DresdenDresdenGermany Qinhuangdao Municipal Bureau of Foreign Affairs and Commerce QinhuangdaoChina

Uncertainty exists in many aspects of building simulation.A deterministic hygrothermal analysis may not sufficiently give a reliable guidance if a number of input variables are subject to *** this paper,a probabilistic-based method was developed to evaluate the hygrothermal performance of building *** approach accounts for the uncertainties from model inputs and propagates them to the outputs through the simulation model,thus it provides a likelihood of performance *** hypercube sampling technique,incorporated with correlation structure among the inputs,was applied to generate the random samples that follows the intrinsic *** performance of an internally insulated masonry wall was evaluated by applying the proposed approach against different *** performance,condensation and mould growth potential of the renovated wall can overall satisfy the requirements stipulated in multifold *** most influential inputs were identified by the standardized regression sensitivity analysis and partial correlation *** methods deliver the same key parameters for the single and time-dependent output variables in the case *** probabilistic method can provide a comprehensive risk analysis and support the decision-maker and engineer in the design and optimization of building components.

关键词： probabilistic approach hygrothermal performance building component uncertainty analysis sensitivity analysis

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Recent behavior and possible future evolution of the glacieret in the cirque Golemiya Kazan in the Pirin Mountains under conditions of climate warming

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Journal of Mountain Science 2019年第1期16卷 16-29页

作者： Peter NOJAROV Emil GACHEV Karsten grunewald National Institute of Geophysics Geodesy and Geography Bulgarian Academy of Sciences South-West University "Neofit Rilski" Leibniz Institute of Ecological Urban and Regional Development

This research reveals relationships between climate variables and inter-annual dynamics in the area of the glacieret located in the cirque Golemiya Kazan in the Pirin Mountains. The study period is 1993–2017. The correlations are identified using statistical methods. Also, a statistical model is constructed, including some climate variables as predictors. Despite the evident decrease of the glacieret's size in the period from the 1950 s onwards, the long-term trends for the last decades have been insignificant. The main climatic factors influencing the inter-annual dynamics in the area of the glacieret are air temperature, precipitation, zonal and meridional winds and relative humidity. With respect to the dynamics in the area of the glacieret, the important trends in the different climate variables are those of the warm period air temperatures and zonal(u) wind. They also determine to a great extent its future development by acting in two opposite directions – rising temperatures in the warm period will lead to a rapid decrease of its area by the end of the melting season, while the change of wind directionfrom west to east in the warm period will increase its area. The influence of the zonal wind in the warm period is explained mainly by the location of the glacieret in the cirque. Generally, the glacieret is tilted downwards from west to east. Thus, westerly winds facilitate blowing away the snow from the surface of the glacieret, assisting its melting in the warm period. Easterly winds do not have such an effect. The combination of the opposite effects of these two most important climate variables leads to the most likely scenario for the future development of the glacieret, according to which by the middle of this century it is expected to turn into a semi-permanent snow patch, which disappears after some summers, and by the end of the century to completely melt every year before the end of the melting season.

关键词： Climate warming Glacieret Pirin Mountains Statistical modeling Glacieret area Projection

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Can DyeCycling break the photobleaching limit in single-molecule FRET?

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Nano Research 2022年第11期15卷 9818-9830页

作者： Benjamin Vermeer sonja Schmid NanoDynamicsLab Laboratory of BiophysicsWageningen UniversityStippeneng 46708WE WageningenThe Netherlands

Biomolecular systems,such as proteins,crucially rely on dynamic processes at the *** biomolecular nanodynamics is therefore key to obtaining a mechanistic understanding of the energies and molecular driving forces that controlbiomolecular ***-molecule fluorescence resonance energy transfer(smFRET)is a powerful technique to observe inreal-time how a single biomolecule proceeds through its functional cycle involving a sequence of distinct structural ***,this technique is fundamentally limited by irreversible photobleaching,causing the untimely end of the experiment andthus,a narrow temporal bandwidth of≤3 orders of ***,we introduce“DyeCycling”,a measurement scheme withwhich we aim to break the photobleaching limit in *** introduce the concept of spontaneous dye replacement bysimulations,and as an experimental proof-of-concept,we demonstrate the intermittent observation of a single biomolecule forone hour with a time resolution of ***,DyeCycling can provide>100-fold more information per singlemolecule than conventional *** discuss the experimental implementation of DyeCycling,its current and fundamentallimitations,and specific biological use *** its general simplicity and versatility,DyeCycling has the potential torevolutionize the field of time-resolved smFRET,where it may serve to unravel a wealth of biomolecular dynamics by bridgingfrom milliseconds to the hour range.

关键词： biomolecular dynamics single-molecule fluorescence resonance energy transfer(smFRET) photobleaching conformational changes single-molecule kinetics

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Preliminary Mapping and Characterisation of Soils in the Pirin Mountains (Bulgaria)

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Journal of Mountain Science 2008年第2期5卷 122-129页

作者： Maika Lβiger Jrg Scheithauer Karsten grunewald University of Technology Dresden D-01062 Dresden Germany Landscape Research Centre Dresden Am Ende 14 D-01277 Dresden Germany

Soil types, humus types and vegetation as well as their hypsometric variation were analysed in terms of sequences in the northern part of the high mountains of the Pirin National Park at altitudes between looo and 2400 m a.s.1. The study area is characterised by a large variety of natural parameters like petrology （mainly marble and granite）, morphology （different slope deposits, exposition） and the orographic climate gradient. Statistical analyses using these parameters provided a basis for the soil group classification of the sites. Based on a Digital Terrain Model （DTM） and a geological map of the Pirin National Park, the results of these statistical analyses were used to generate a ＂map of potential soil groups＂（regionalisation using GIS）. Six potential soil groups could be determined. The resulting map exhibits a confidence level of 68 % on 74.4 % of the covered area. Rendzic Leptosols, in combination with Folic Histosols and Histi-lithic Leptosols occur in the alpine and subalpine regions on calcareous substrates. With decreasing altitude they are replaced by a mosaic of Rendzic Leptosols, Phaeozems and an increasing occurrence of Cambic Umbrisols. Umbrisols found on silicatic substrates in the alpine region are replaced by Cambic Umbrisols with decreasing altitude as well. Hence, pedogenesis is characterised by increasing browning and depth of the soil profiles with decreasing altitude. The pH-level is slightly acidic to neutral in lower zones and on calcareous rocky bases. Acidification increases in the subalpine zone. Soil pH decreases down to 4 on silicate subtrates. Typical humic values in mineral topsoils are 10 to 12 %, and in organic layers of the soils above 2000 m a.s.1, they are even more. The C：N ratio closely ranges around 20 （median）.

关键词： Cluster analysis soil mapping hypsometric soil differentiation Southeast Europe

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Optogenetic neuroregeneration

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Neural Regeneration Research 2022年第7期17卷 1468-1470页

作者： Harald Janovjak sonja Kleinlogel EMBL Australia Australian Regenerative Medicine InstituteFaculty of MedicineNursing and Health SciencesMonash UniversityClayton/MelbourneAustralia Institute of Physiology University of BernBernSwitzerland

Optogenetics is a powerful technology that employs light and genetics to manipulate physiology and behavior with unprecedented *** high acuity of light stimulation permits fine control both in space(e.g.,to target just one tissue in an animal)and in time(e.g.,to interfere with a specific disease stage),whilst genetic targeting restricts manipulation to a functionally-relevant cell population(Figure 1A).

关键词： stimulation manipulation Figure

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