Publications by authors named "Albrecht Röpke"

46 Publications

The first versatile human iPSC-based model of ectopic virus induction allows new insights in RNA-virus disease.

Sci Rep 2020 10 8;10(1):16804. Epub 2020 Oct 8.

Institute for Genetics of Heart Diseases (IfGH), Department of Cardiovascular Medicine, University Hospital Münster, 48149, Münster, Germany.

A detailed description of pathophysiological effects that viruses exert on their host is still challenging. For the first time, we report a highly controllable viral expression model based on an iPS-cell line from a healthy human donor. The established viral model system enables a dose-dependent and highly localized RNA-virus expression in a fully controllable environment, giving rise for new applications for the scientific community.
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http://dx.doi.org/10.1038/s41598-020-72966-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7546621PMC
October 2020

Generation of a human iPSC line (MPIi007-A) from a patient with Metachromatic leukodystrophy.

Stem Cell Res 2020 10 13;48:101993. Epub 2020 Sep 13.

Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Röntgenstrasse 20, Münster 48149, Germany; Medical Faculty, University of Münster, Domagkstrasse 3, Münster 48149, Germany. Electronic address:

Here we have generated a human induced pluripotent stem cells (hiPSC) line (MPIi007-A) from skin fibroblasts of a 4-year-old male Metachromatic leukodystrophy (MLD) patient with a heterozygous 1178C > G (Thr393Ser) mutation in arylsulfatase A (ARSA) gene via retroviral expression of OCT4, SOX2, KLF4 and c-MYC. The MPIi007-A iPSC line displayed typical embryonic stem cell-like morphology, carried the ARSA gene mutation, expressed several pluripotent stem cell makers, retained normal karyotype (46, XY) and were capable of forming teratomas containing three germ layers. The MPIi007-A line can be used for the characterization of MLD-associated pathomechanisms and developing new therapeutic options.
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http://dx.doi.org/10.1016/j.scr.2020.101993DOI Listing
October 2020

Reprogramming competence of OCT factors is determined by transactivation domains.

Sci Adv 2020 Sep 2;6(36). Epub 2020 Sep 2.

Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Röntgenstrasse 20, Münster 48149, Germany.

OCT4 (also known as POU5F1) plays an essential role in reprogramming. It is the only member of the POU (Pit-Oct-Unc) family of transcription factors that can induce pluripotency despite sharing high structural similarities to all other members. Here, we discover that OCT6 (also known as POU3F1) can elicit reprogramming specifically in human cells. OCT6-based reprogramming does not alter the mesenchymal-epithelial transition but is attenuated through the delayed activation of the pluripotency network in comparison with OCT4-based reprogramming. Creating a series of reciprocal domain-swapped chimeras and mutants across all OCT factors, we clearly delineate essential elements of OCT4/OCT6-dependent reprogramming and, conversely, identify the features that prevent induction of pluripotency by other OCT factors. With this strategy, we further discover various chimeric proteins that are superior to OCT4 in reprogramming. Our findings clarify how reprogramming competences of OCT factors are conferred through their structural components.
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http://dx.doi.org/10.1126/sciadv.aaz7364DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7467702PMC
September 2020

Extrinsic immune cell-derived, but not intrinsic oligodendroglial factors contribute to oligodendroglial differentiation block in multiple sclerosis.

Acta Neuropathol 2020 11 7;140(5):715-736. Epub 2020 Sep 7.

Institute of Neuropathology, University Hospital Münster, Pottkamp 2, 48149, Münster, Germany.

Multiple sclerosis (MS) is the most frequent demyelinating disease in young adults and despite significant advances in immunotherapy, disease progression still cannot be prevented. Promotion of remyelination, an endogenous repair mechanism resulting in the formation of new myelin sheaths around demyelinated axons, represents a promising new treatment approach. However, remyelination frequently fails in MS lesions, which can in part be attributed to impaired differentiation of oligodendroglial progenitor cells into mature, myelinating oligodendrocytes. The reasons for impaired oligodendroglial differentiation and defective remyelination in MS are currently unknown. To determine whether intrinsic oligodendroglial factors contribute to impaired remyelination in relapsing-remitting MS (RRMS), we compared induced pluripotent stem cell-derived oligodendrocytes (hiOL) from RRMS patients and controls, among them two monozygous twin pairs discordant for MS. We found that hiOL from RRMS patients and controls were virtually indistinguishable with respect to remyelination-associated functions and proteomic composition. However, while analyzing the effect of extrinsic factors we discovered that supernatants of activated peripheral blood mononuclear cells (PBMCs) significantly inhibit oligodendroglial differentiation. In particular, we identified CD4 T cells as mediators of impaired oligodendroglial differentiation; at least partly due to interferon-gamma secretion. Additionally, we observed that blocked oligodendroglial differentiation induced by PBMC supernatants could not be restored by application of oligodendroglial differentiation promoting drugs, whereas treatment of PBMCs with the immunomodulatory drug teriflunomide prior to supernatant collection partly rescued oligodendroglial differentiation. In summary, these data indicate that the oligodendroglial differentiation block is not due to intrinsic oligodendroglial factors but rather caused by the inflammatory environment in RRMS lesions which underlines the need for drug screening approaches taking the inflammatory environment into account. Combined, these findings may contribute to the development of new remyelination promoting strategies.
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http://dx.doi.org/10.1007/s00401-020-02217-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7547031PMC
November 2020

The Ca channel CatSper is not activated by cAMP/PKA signaling but directly affected by chemicals used to probe the action of cAMP and PKA.

J Biol Chem 2020 09 23;295(38):13181-13193. Epub 2020 Jul 23.

Centre of Reproductive Medicine and Andrology, University Hospital Münster, University of Münster, Münster, Germany; Cells in Motion Interfaculty Centre, University of Münster, Münster, Germany. Electronic address:

The sperm-specific Ca channel CatSper (cation channel of sperm) controls the influx of Ca into the flagellum and, thereby, the swimming behavior of sperm. A hallmark of human CatSper is its polymodal activation by membrane voltage, intracellular pH, and oviductal hormones. Whether CatSper is also activated by signaling pathways involving an increase of cAMP and ensuing activation of PKA is, however, a matter of controversy. To shed light on this question, we used kinetic ion-sensitive fluorometry, patch-clamp recordings, and optochemistry to study transmembrane Ca flux and membrane currents in human sperm from healthy donors and from patients that lack functional CatSper channels. We found that human CatSper is neither activated by intracellular cAMP directly nor indirectly by the cAMP/PKA-signaling pathway. Instead, we show that nonphysiological concentrations of cAMP and membrane-permeable cAMP analogs used to mimic the action of intracellular cAMP activate human CatSper from the outside via a hitherto-unknown extracellular binding site. Finally, we demonstrate that the effects of common PKA inhibitors on human CatSper rest predominantly, if not exclusively, on off-target drug actions on CatSper itself rather than on inhibition of PKA. We conclude that the concept of an intracellular cAMP/PKA-activation of CatSper is primarily based on unspecific effects of chemical probes used to interfere with cAMP signaling. Altogether, our findings solve several controversial issues and reveal a novel ligand-binding site controlling the activity of CatSper, which has important bearings on future studies of cAMP and Ca signaling in sperm.
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http://dx.doi.org/10.1074/jbc.RA120.013218DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7504923PMC
September 2020

Bi-allelic Mutations in M1AP Are a Frequent Cause of Meiotic Arrest and Severely Impaired Spermatogenesis Leading to Male Infertility.

Am J Hum Genet 2020 08 15;107(2):342-351. Epub 2020 Jul 15.

Institute of Human Genetics, University of Münster, 48149 Münster, Germany. Electronic address:

Male infertility affects ∼7% of men, but its causes remain poorly understood. The most severe form is non-obstructive azoospermia (NOA), which is, in part, caused by an arrest at meiosis. So far, only a few validated disease-associated genes have been reported. To address this gap, we performed whole-exome sequencing in 58 men with unexplained meiotic arrest and identified the same homozygous frameshift variant c.676dup (p.Trp226LeufsTer4) in M1AP, encoding meiosis 1 associated protein, in three unrelated men. This variant most likely results in a truncated protein as shown in vitro by heterologous expression of mutant M1AP. Next, we screened four large cohorts of infertile men and identified three additional individuals carrying homozygous c.676dup and three carrying combinations of this and other likely causal variants in M1AP. Moreover, a homozygous missense variant, c.1166C>T (p.Pro389Leu), segregated with infertility in five men from a consanguineous Turkish family. The common phenotype between all affected men was NOA, but occasionally spermatids and rarely a few spermatozoa in the semen were observed. A similar phenotype has been described for mice with disruption of M1ap. Collectively, these findings demonstrate that mutations in M1AP are a relatively frequent cause of autosomal recessive severe spermatogenic failure and male infertility with strong clinical validity.
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http://dx.doi.org/10.1016/j.ajhg.2020.06.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7413853PMC
August 2020

Crisponi syndrome/cold-induced sweating syndrome type 2: Reprogramming of CS/CISS2 individual derived fibroblasts into three clones of one iPSC line.

Stem Cell Res 2020 07 1;46:101855. Epub 2020 Jun 1.

Department of General Pediatrics, Muenster University Children's Hospital, Muenster, Germany.

Crisponi syndrome/cold-induced sweating syndrome type 2 (CS/CISS2) is a rare disease with severe dysfunctions of thermoregulatory processes. CS/CISS2 individuals suffer from recurrent episodes of hyperthermia in the neonatal period and paradoxical sweating at cold ambient temperatures in adolescence. Variants in CLCF1 (cardiotrophin-like-cytokine 1) cause CS/CISS2. Here, we summarize the generation of three clones of one stem cell line (iPSC) of a CS/CISS2 individual carrying the CLCF1 variant c.321C>G on both alleles. These patient derived iPSC clones show a normal karyotype, several pluripotency markers, and the ability to differentiate into the three germ layers.
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http://dx.doi.org/10.1016/j.scr.2020.101855DOI Listing
July 2020

Generation of induced pluripotent stem cell lines from a Crisponi/Cold induced sweating syndrome type 1 individual.

Stem Cell Res 2020 07 28;46:101820. Epub 2020 May 28.

Department of General Pediatrics, Muenster University Children's Hospital, Muenster, Germany.

Cytokine receptor like factor 1 (CRLF1) is the gene implicated, when mutated, in Crisponi syndrome/cold-induced sweating syndrome type 1 (CS/CISS1). Here, we report the establishment of induced pluripotent stem cell lines (iPSCs) from fibroblasts of a Turkish CS/CISS1 individual with a homozygous variant in CRLF1 (c.708_709delinsT; p.[Pro238Argfs*6]). This variant is the most frequent variant associated to CS/CISS1 in the Turkish population. These patient derived iPSC lines show all pluripotency markers, a normal karyotype and the ability to differentiate into the three germ layers.
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http://dx.doi.org/10.1016/j.scr.2020.101820DOI Listing
July 2020

Generation of a human iPSC line (MPIi006-A) from a patient with Pelizaeus-Merzbacher disease.

Stem Cell Res 2020 07 7;46:101839. Epub 2020 May 7.

Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Röntgenstrasse 20, Münster 48149, Germany; Medical Faculty, University of Münster, Domagkstrasse 3, Münster 48149, Germany. Electronic address:

We established a human induced pluripotent stem cells (hiPSC) line (MPIi006-A) from fibroblasts of a 20-year-old male Pelizaeus-Merzbacher disease (PMD) patient with a hemizygous 643C>T mutation in proteolipid protein 1 (PLP1) gene using a retroviral delivery of OCT4, SOX2, KLF4 and c-MYC. The MPIi006-A iPSC line carried the mutation, displayed typical iPSC morphology, expressed pluripotent stem cell makers, exhibited normal karyotype and were capable of differentiating into cells representative of three germ layers.
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http://dx.doi.org/10.1016/j.scr.2020.101839DOI Listing
July 2020

Initial experience with [F]DPA-714 TSPO-PET to image inflammation in primary angiitis of the central nervous system.

Eur J Nucl Med Mol Imaging 2020 08 20;47(9):2131-2141. Epub 2020 Jan 20.

Department of Neurology with Institute of Translational Neurology, University Hospital Münster, Münster, Germany.

Purpose: Primary angiitis of the central nervous system (PACNS) is a heterogeneous, rare, and poorly understood inflammatory disease. We aimed at non-invasive imaging of activated microglia/macrophages in patients with PACNS by PET-MRI targeting the translocator protein (TSPO) with [F]DPA-714 to potentially assist differential diagnosis, therapy monitoring, and biopsy planning.

Methods: In total, nine patients with ischemic stroke and diagnosed or suspected PACNS underwent [F]DPA-714-PET-MRI. Dynamic PET scanning was performed for 60 min after injection of 233 ± 19 MBq [F]DPA-714, and MRI was simultaneously acquired.

Results: In two PACNS patients, [F]DPA-714 uptake patterns exceeded MRI correlates of infarction, whereas uptake was confined to the infarct in four patients where initial suspicion of PACNS could not be confirmed. About three patients with PACNS or cerebral predominant lymphocytic vasculitis showed no or only faintly increased uptake. Short-term [F]DPA-714-PET follow-up in a patient with PACNS showed reduced lesional [F]DPA-714 uptake after anti-inflammatory treatment. Biopsy in the same patient pinpointed the source of tracer uptake to TSPO-expressing immune cells.

Conclusions: [F]DPA-714-PET imaging may facilitate the diagnosis and treatment monitoring of PACNS. Further studies are needed to fully understand the potential of TSPO-PET in deciphering the heterogeneity of the disease.
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http://dx.doi.org/10.1007/s00259-019-04662-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7338821PMC
August 2020

Rotational motion and rheotaxis of human sperm do not require functional CatSper channels and transmembrane Ca signaling.

EMBO J 2020 02 19;39(4):e102363. Epub 2020 Jan 19.

Centre of Reproductive Medicine and Andrology, University Hospital Münster, University of Münster, Münster, Germany.

Navigation of sperm in fluid flow, called rheotaxis, provides long-range guidance in the mammalian oviduct. The rotation of sperm around their longitudinal axis (rolling) promotes rheotaxis. Whether sperm rolling and rheotaxis require calcium (Ca ) influx via the sperm-specific Ca channel CatSper, or rather represent passive biomechanical and hydrodynamic processes, has remained controversial. Here, we study the swimming behavior of sperm from healthy donors and from infertile patients that lack functional CatSper channels, using dark-field microscopy, optical tweezers, and microfluidics. We demonstrate that rolling and rheotaxis persist in CatSper-deficient human sperm. Furthermore, human sperm undergo rolling and rheotaxis even when Ca influx is prevented. Finally, we show that rolling and rheotaxis also persist in mouse sperm deficient in both CatSper and flagellar Ca -signaling domains. Our results strongly support the concept that passive biomechanical and hydrodynamic processes enable sperm rolling and rheotaxis, rather than calcium signaling mediated by CatSper or other mechanisms controlling transmembrane Ca flux.
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http://dx.doi.org/10.15252/embj.2019102363DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7024840PMC
February 2020

Disorders of spermatogenesis: Perspectives for novel genetic diagnostics after 20 years of unchanged routine.

Med Genet 2018 26;30(1):12-20. Epub 2018 Feb 26.

Institute of Human Genetics, University of Münster, Vesaliusweg 12-14, 48149 Münster, Germany.

Infertility is a common condition estimated to affect 10-15% of couples. The clinical causes are attributed in equal parts to the male and female partners. Diagnosing male infertility mostly relies on semen (and hormone) analysis, which results in classification into the two major phenotypes of oligo- and azoospermia. The clinical routine analyses have not changed over the last 20 years and comprise screening for chromosomal aberrations and Y‑chromosomal azoospermia factor deletions. These tests establish a causal genetic diagnosis in about 4% of unselected men in infertile couples and 20% of azoospermic men. Gene sequencing is currently only performed in very rare cases of hypogonadotropic hypogonadism and the gene is routinely analysed in men with obstructive azoospermia. Still, a large number of genes have been proposed to be associated with male infertility by, for example, knock-out mouse models. In particular, those that are exclusively expressed in the testes are potential candidates for further analyses. However, the genome-wide analyses (a few array-CGH, six GWAS, and some small exome sequencing studies) performed so far have not lead to improved clinical diagnostic testing. In 2017, we started to routinely analyse the three validated male infertility genes: , and . Preliminary analyses demonstrated highly likely pathogenic mutations in these genes as a cause of azoospermia in 4 men, equalling 5% of the 80 patients analysed so far, and increasing the diagnostic yield in this group to 25%. Over the past few years, we have observed a steep increase in publications on novel candidate genes for male infertility, especially in men with azoospermia. In addition, concerted efforts to achieve progress in elucidating genetic causes of male infertility and to introduce novel testing strategies into clinical routine have been made recently. Thus, we are confident that major breakthroughs concerning the genetics of male infertility will be achieved in the near future and will translate into clinical routine to improve patient/couple care.
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http://dx.doi.org/10.1007/s11825-018-0181-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5838132PMC
February 2018

Cardiogenic programming of human pluripotent stem cells by dose-controlled activation of EOMES.

Nat Commun 2018 01 30;9(1):440. Epub 2018 Jan 30.

Human Stem Cell Pluripotency Laboratory, Max Planck Institute for Molecular Biomedicine, 48149, Münster, Germany.

Master cell fate determinants are thought to induce specific cell lineages in gastrulation by orchestrating entire gene programs. The T-box transcription factor EOMES (eomesodermin) is crucially required for the development of the heart-yet it is equally important for endoderm specification suggesting that it may act in a context-dependent manner. Here, we define an unrecognized interplay between EOMES and the WNT signaling pathway in controlling cardiac induction by using loss and gain-of-function approaches in human embryonic stem cells. Dose-dependent EOMES induction alone can fully replace a cocktail of signaling molecules otherwise essential for the specification of cardiogenic mesoderm. Highly efficient cardiomyocyte programming by EOMES mechanistically involves autocrine activation of canonical WNT signaling via the WNT3 ligand, which necessitates a shutdown of this axis at a subsequent stage. Our findings provide insights into human germ layer induction and bear biotechnological potential for the robust production of cardiomyocytes from engineered stem cells.
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http://dx.doi.org/10.1038/s41467-017-02812-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5789885PMC
January 2018

Action of steroids and plant triterpenoids on CatSper Ca channels in human sperm.

Proc Natl Acad Sci U S A 2018 01 5;115(3):E344-E346. Epub 2018 Jan 5.

Center of Reproductive Medicine and Andrology, University Hospital Münster, 48149 Münster, Germany;

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http://dx.doi.org/10.1073/pnas.1717929115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5777000PMC
January 2018

Cardiac Subtype-Specific Modeling of K1.5 Ion Channel Deficiency Using Human Pluripotent Stem Cells.

Front Physiol 2017 6;8:469. Epub 2017 Jul 6.

Human Stem Cell Pluripotency Laboratory, Max Planck Institute for Molecular BiomedicineMünster, Germany.

The ultrarapid delayed rectifier K current (I), mediated by K1.5 channels, constitutes a key component of the atrial action potential. Functional mutations in the underlying gene have been shown to cause hereditary forms of atrial fibrillation (AF). Here, we combine targeted genetic engineering with cardiac subtype-specific differentiation of human induced pluripotent stem cells (hiPSCs) to explore the role of K1.5 in atrial hiPSC-cardiomyocytes. CRISPR/Cas9-mediated mutagenesis of integration-free hiPSCs was employed to generate a functional knockout. This model as well as isogenic wild-type control hiPSCs could selectively be differentiated into ventricular or atrial cardiomyocytes at high efficiency, based on the specific manipulation of retinoic acid signaling. Investigation of electrophysiological properties in K1.5-deficient cardiomyocytes compared to isogenic controls revealed a strictly atrial-specific disease phentoype, characterized by cardiac subtype-specific field and action potential prolongation and loss of 4-aminopyridine sensitivity. Atrial K1.5-deficient cardiomyocytes did not show signs of arrhythmia under adrenergic stress conditions or upon inhibiting additional types of K current. Exposure of bulk cultures to carbachol lowered beating frequencies and promoted chaotic spontaneous beating in a stochastic manner. Low-frequency, electrical stimulation in single cells caused atrial and mutant-specific early afterdepolarizations, linking the loss of function to a putative trigger mechanism in familial AF. These results clarify for the first time the role of K1.5 in atrial hiPSC-cardiomyocytes and demonstrate the feasibility of cardiac subtype-specific disease modeling using engineered hiPSCs.
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http://dx.doi.org/10.3389/fphys.2017.00469DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5498524PMC
July 2017

Generation and cardiac subtype-specific differentiation of PITX2-deficient human iPS cell lines for exploring familial atrial fibrillation.

Stem Cell Res 2017 05 23;21:26-28. Epub 2017 Mar 23.

Human Stem Cell Pluripotency Laboratory, Max Planck Institute for Molecular Biomedicine, Münster, Germany; Chemical Genomics Centre of the Max Planck Society, Dortmund, Germany. Electronic address:

Loss-of-function mutations in the PITX2 transcription factor gene have been shown to cause familial atrial fibrillation (AF). To potentially model aspects of AF and unravel PITX2-regulated downstream genes for drug target discovery, we here report the generation of integration-free PITX2-deficient hiPS cell lines. We also show that both PITX2 knockout hiPS cells and isogenic wild-type controls can selectively be differentiated into human atrial cardiomyocytes, to potentially uncover differentially expressed gene sets between these groups.
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http://dx.doi.org/10.1016/j.scr.2017.03.015DOI Listing
May 2017

MECHANISMS IN ENDOCRINOLOGY: Aberrations of the X chromosome as cause of male infertility.

Eur J Endocrinol 2017 Nov 13;177(5):R249-R259. Epub 2017 Jun 13.

Male infertility is most commonly caused by spermatogenetic failure, clinically noted as oligo- or a-zoospermia. Today, in approximately 20% of azoospermic patients, a causal genetic defect can be identified. The most frequent genetic causes of azoospermia (or severe oligozoospermia) are Klinefelter syndrome (47,XXY), structural chromosomal abnormalities and Y-chromosomal microdeletions. Consistent with Ohno's law, the human X chromosome is the most stable of all the chromosomes, but contrary to Ohno's law, the X chromosome is loaded with regions of acquired, rapidly evolving genes, which are of special interest because they are predominantly expressed in the testis. Therefore, it is not surprising that the X chromosome, considered as the female counterpart of the male-associated Y chromosome, may actually play an essential role in male infertility and sperm production. This is supported by the recent description of a significantly increased copy number variation (CNV) burden on both sex chromosomes in infertile men and point mutations in X-chromosomal genes responsible for male infertility. Thus, the X chromosome seems to be frequently affected in infertile male patients. Four principal X-chromosomal aberrations have been identified so far: (1) aneuploidy of the X chromosome as found in Klinefelter syndrome (47,XXY or mosaicism for additional X chromosomes). (2) Translocations involving the X chromosome, e.g. nonsyndromic 46,XX testicular disorders of sex development (XX-male syndrome) or X-autosome translocations. (3) CNVs affecting the X chromosome. (4) Point mutations disrupting X-chromosomal genes. All these are reviewed herein and assessed concerning their importance for the clinical routine diagnostic workup of the infertile male as well as their potential to shape research on spermatogenic failure in the next years.
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http://dx.doi.org/10.1530/EJE-17-0246DOI Listing
November 2017

A no-stop mutation in MAGEB4 is a possible cause of rare X-linked azoospermia and oligozoospermia in a consanguineous Turkish family.

J Assist Reprod Genet 2017 May 11;34(5):683-694. Epub 2017 Apr 11.

Département Génomique Fonctionnelle et Cancer, Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Institut National de Santé et de Recherche Médicale (INSERM) U964/Centre National de Recherche Scientifique (CNRS) UMR 7104, Université de Strasbourg, 67404, Illkirch, France.

Purpose: The purpose of this study was to identify mutations that cause non-syndromic male infertility using whole exome sequencing of family cases.

Methods: We recruited a consanguineous Turkish family comprising nine siblings with male triplets; two of the triplets were infertile as well as one younger infertile brother. Whole exome sequencing (WES) performed on two azoospermic brothers identified a mutation in the melanoma antigen family B4 (MAGEB4) gene which was confirmed via Sanger sequencing and then screened for on control groups and unrelated infertile subjects. The effect of the mutation on messenger RNA (mRNA) and protein levels was tested after in vitro cell transfection. Structural features of MAGEB4 were predicted throughout the conserved MAGE domain.

Results: The novel single-base substitution (c.1041A>T) in the X-linked MAGEB4 gene was identified as a no-stop mutation. The mutation is predicted to add 24 amino acids to the C-terminus of MAGEB4. Our functional studies were unable to detect any effect either on mRNA stability, intracellular localization of the protein, or the ability to homodimerize/heterodimerize with other MAGE proteins. We thus hypothesize that these additional amino acids may affect the proper protein interactions with MAGEB4 partners.

Conclusion: The whole exome analysis of a consanguineous Turkish family revealed MAGEB4 as a possible new X-linked cause of inherited male infertility. This study provides the first clue to the physiological function of a MAGE protein.
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http://dx.doi.org/10.1007/s10815-017-0900-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5427651PMC
May 2017

Astrocyte pathology in a human neural stem cell model of frontotemporal dementia caused by mutant TAU protein.

Sci Rep 2017 03 3;7:42991. Epub 2017 Mar 3.

Max Planck Institute for Molecular Biomedicine, Department of Cell and Developmental Biology, 48149 Münster, Germany.

Astroglial pathology is seen in various neurodegenerative diseases including frontotemporal dementia (FTD), which can be caused by mutations in the gene encoding the microtubule-associated protein TAU (MAPT). Here, we applied a stem cell model of FTD to examine if FTD astrocytes carry an intrinsic propensity to degeneration and to determine if they can induce non-cell-autonomous effects in neighboring neurons. We utilized CRISPR/Cas9 genome editing in human induced pluripotent stem (iPS) cell-derived neural progenitor cells (NPCs) to repair the FTD-associated N279K MAPT mutation. While astrocytic differentiation was not impaired in FTD NPCs derived from one patient carrying the N279K MAPT mutation, FTD astrocytes appeared larger, expressed increased levels of 4R-TAU isoforms, demonstrated increased vulnerability to oxidative stress and elevated protein ubiquitination and exhibited disease-associated changes in transcriptome profiles when compared to astrocytes derived from one control individual and to the isogenic control. Interestingly, co-culture experiments with FTD astrocytes revealed increased oxidative stress and robust changes in whole genome expression in previously healthy neurons. Our study highlights the utility of iPS cell-derived NPCs to elucidate the role of astrocytes in the pathogenesis of FTD.
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http://dx.doi.org/10.1038/srep42991DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5335603PMC
March 2017

The human RHOX gene cluster: target genes and functional analysis of gene variants in infertile men.

Hum Mol Genet 2016 11;25(22):4898-4910

Centre of Reproductive Medicine and Andrology, University of Münster, Münster, Germany.

The X-linked reproductive homeobox (RHOX) gene cluster encodes transcription factors preferentially expressed in reproductive tissues. This gene cluster has important roles in male fertility based on phenotypic defects of Rhox-mutant mice and the finding that aberrant RHOX promoter methylation is strongly associated with abnormal human sperm parameters. However, little is known about the molecular mechanism of RHOX function in humans. Using gene expression profiling, we identified genes regulated by members of the human RHOX gene cluster. Some genes were uniquely regulated by RHOXF1 or RHOXF2/2B, while others were regulated by both of these transcription factors. Several of these regulated genes encode proteins involved in processes relevant to spermatogenesis; e.g. stress protection and cell survival. One of the target genes of RHOXF2/2B is RHOXF1, suggesting cross-regulation to enhance transcriptional responses. The potential role of RHOX in human infertility was addressed by sequencing all RHOX exons in a group of 250 patients with severe oligozoospermia. This revealed two mutations in RHOXF1 (c.515G > A and c.522C > T) and four in RHOXF2/2B (-73C > G, c.202G > A, c.411C > T and c.679G > A), of which only one (c.202G > A) was found in a control group of men with normal sperm concentration. Functional analysis demonstrated that c.202G > A and c.679G > A significantly impaired the ability of RHOXF2/2B to regulate downstream genes. Molecular modelling suggested that these mutations alter RHOXF2/F2B protein conformation. By combining clinical data with in vitro functional analysis, we demonstrate how the X-linked RHOX gene cluster may function in normal human spermatogenesis and we provide evidence that it is impaired in human male fertility.
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http://dx.doi.org/10.1093/hmg/ddw313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6281360PMC
November 2016

Intrafamilial phenotypic variability of Specific Language Impairment.

Brain Lang 2016 Aug 4;159:102-8. Epub 2016 Jul 4.

Department of Phoniatrics and Pedaudiology, University Hospital of Muenster, Kardinal-von-Galen Ring 10, 48149 Muenster, Germany. Electronic address:

We investigated language functions in 32 members of a four generation family with several members affected by Specific Language Impairment with an extensive language test battery in order to determine the prevalence, overlap, and homogeneity of linguistic deficits within one pedigree. In sum, one fourth of all family members tested fulfilled the criteria of Specific Language Impairment. Despite of some similarities in language abilities, different combinations of language deficits were observed, and individual language profiles varied substantially. Thus, though there is a high prevalence of language deficits in this family which raises the likelihood of a genetic origin of these deficits, and though all affected study participants displayed selective linguistic deficits with normal non-verbal functioning, language testing showed considerable variance in overlap and homogeneity of linguistic deficits. Thus, even in one genetic population, an underlying linguistic disorder manifests itself in different language abilities to a variant degree.
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http://dx.doi.org/10.1016/j.bandl.2016.06.005DOI Listing
August 2016

Distinct Neurodegenerative Changes in an Induced Pluripotent Stem Cell Model of Frontotemporal Dementia Linked to Mutant TAU Protein.

Stem Cell Reports 2015 Jul 2;5(1):83-96. Epub 2015 Jul 2.

Max Planck Institute for Molecular Biomedicine, 48149 Münster, Germany; Institute of Neuropathology, University Hospital Münster, 48149 Münster, Germany. Electronic address:

Frontotemporal dementia (FTD) is a frequent form of early-onset dementia and can be caused by mutations in MAPT encoding the microtubule-associated protein TAU. Because of limited availability of neural cells from patients' brains, the underlying mechanisms of neurodegeneration in FTD are poorly understood. Here, we derived induced pluripotent stem cells (iPSCs) from individuals with FTD-associated MAPT mutations and differentiated them into mature neurons. Patient iPSC-derived neurons demonstrated pronounced TAU pathology with increased fragmentation and phospho-TAU immunoreactivity, decreased neurite extension, and increased but reversible oxidative stress response to inhibition of mitochondrial respiration. Furthermore, FTD neurons showed an activation of the unfolded protein response, and a transcriptome analysis demonstrated distinct, disease-associated gene expression profiles. These findings indicate distinct neurodegenerative changes in FTD caused by mutant TAU and highlight the unique opportunity to use neurons differentiated from patient-specific iPSCs to identify potential targets for drug screening purposes and therapeutic intervention.
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http://dx.doi.org/10.1016/j.stemcr.2015.06.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4618448PMC
July 2015

X-linked TEX11 mutations, meiotic arrest, and azoospermia in infertile men.

N Engl J Med 2015 May 13;372(22):2097-107. Epub 2015 May 13.

From the Departments of Obstetrics, Gynecology, and Reproductive Sciences (A.N.Y., A.P.G., J.S., A. Rajkovic, S.A.Y.) and Urology (T.J.), University of Pittsburgh School of Medicine, and the Department of Biological Sciences, University of Pittsburgh, Kenneth P. Dietrich School of Arts and Sciences (A.J.B.) - all in Pittsburgh; the Institute of Human Genetics (A. Röpke, F.T.) and Center of Reproductive Medicine and Andrology (B.W., S.K., S.S.), University of Münster, Münster, Germany; and the Department of Reproductive Biology and Stem Cells, Institute of Human Genetics, Polish Academy of Sciences, Poznań (M.O., M.K.).

Background: The genetic basis of nonobstructive azoospermia is unknown in the majority of infertile men.

Methods: We performed array comparative genomic hybridization testing in blood samples obtained from 15 patients with azoospermia, and we performed mutation screening by means of direct Sanger sequencing of the testis-expressed 11 gene (TEX11) open reading frame in blood and semen samples obtained from 289 patients with azoospermia and 384 controls.

Results: We identified a 99-kb hemizygous loss on chromosome Xq13.2 that involved three TEX11 exons. This loss, which was identical in 2 patients with azoospermia, predicts a deletion of 79 amino acids within the meiosis-specific sporulation domain SPO22. Our subsequent mutation screening showed five novel TEX11 mutations: three splicing mutations and two missense mutations. These mutations, which occurred in 7 of 289 men with azoospermia (2.4%), were absent in 384 controls with normal sperm concentrations (P=0.003). Notably, five of those TEX11 mutations were detected in 33 patients (15%) with azoospermia who received a diagnosis of azoospermia with meiotic arrest. Meiotic arrest in these patients resembled the phenotype of Tex11-deficient male mice. Immunohistochemical analysis showed specific cytoplasmic TEX11 expression in late spermatocytes, as well as in round and elongated spermatids, in normal human testes. In contrast, testes of patients who had azoospermia with TEX11 mutations had meiotic arrest and lacked TEX11 expression.

Conclusions: In our study, hemizygous TEX11 mutations were a common cause of meiotic arrest and azoospermia in infertile men. (Funded by the National Institutes of Health and others.).
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http://dx.doi.org/10.1056/NEJMoa1406192DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4470617PMC
May 2015

Genetic Variants of DICE1/INTS6 in German Prostate Cancer Families with Linkage to 13q14.

Urol Int 2015 31;95(4):386-9. Epub 2015 Jan 31.

Klinik fx00FC;r Urologie, Otto-von-Guericke-Universitx00E4;t, Magdeburg, Germany.

Introduction: Prostate cancer is the most frequent malignancy found to occur in Caucasian men, but its genetic basis remains elusive. A prostate cancer-susceptibility locus has been identified on chromosome 13q14. The tumour suppressor gene deleted in cancer cells 1 (DICE1/INTS6) is located within this interval on 13q14.3.

Materials And Methods: We performed mutation analysis of the DICE1/INTS6 gene in thirteen German prostate cancer families.

Results And Conclusion: None of the patients harboured DICE1 mutations, and similar frequencies of the previously identified 13 bp deletion polymorphism in the DICE1 promoter were observed in the familial prostate cancer patients as compared with sporadic prostate cancer patients and controls. However, in one family with three affected brothers, the variations c.1215A>C (p.T405T) in exon 10 and c.2568A>G (p.S856S) in exon 17 were detected in a heterozygous pattern. In sporadic prostate cancer patients, variant c.2568A>G (p.S856S) was detected in 10/325 (3.08%) compared with 5/207 (2.42%) control samples (p > 0.05). We conclude that DICE1 appears to be involved in prostate cancer progression rather than in the initiation of prostate cancer.
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http://dx.doi.org/10.1159/000366229DOI Listing
September 2016

Universal cardiac induction of human pluripotent stem cells in two and three-dimensional formats: implications for in vitro maturation.

Stem Cells 2015 May;33(5):1456-69

Human Stem Cell Pluripotency Group; Chemical Genomics Centre of the Max Planck Society, Dortmund, Germany.

Directed cardiac differentiation of human pluripotent stem cells (hPSCs) enables disease modeling, investigation of human cardiogenesis, as well as large-scale production of cardiomyocytes (CMs) for translational purposes. Multiple CM differentiation protocols have been developed to individually address specific requirements of these diverse applications, such as enhanced purity at a small scale or mass production at a larger scale. However, there is no universal high-efficiency procedure for generating CMs both in two-dimensional (2D) and three-dimensional (3D) culture formats, and undefined or complex media additives compromise functional analysis or cost-efficient upscaling. Using systematic combinatorial optimization, we have narrowed down the key requirements for efficient cardiac induction of hPSCs. This implied differentiation in simple serum and serum albumin-free basal media, mediated by a minimal set of signaling pathway manipulations at moderate factor concentrations. The method was applicable both to 2D and 3D culture formats as well as to independent hPSC lines. Global time-course gene expression analyses over extended time periods and in comparison with human heart tissue were used to monitor culture-induced maturation of the resulting CMs. This suggested that hPSC-CMs obtained with our procedure reach a rather stable transcriptomic state after approximately 4 weeks of culture. The underlying gene expression changes correlated well with a decline of immature characteristics as well as with a gain of structural and physiological maturation features within this time frame. These data link gene expression patterns of hPSC-CMs to functional readouts and thus define the cornerstones of culture-induced maturation.
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http://dx.doi.org/10.1002/stem.1964DOI Listing
May 2015

Azoospermia and ring chromosome 9--a case report.

J Assist Reprod Genet 2015 Feb 2;32(2):293-6. Epub 2014 Dec 2.

The Fertility Clinic, Skive Regional Hospital, Skive, Denmark,

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http://dx.doi.org/10.1007/s10815-014-0388-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4354179PMC
February 2015

Mosaicism for an unbalanced Y;21 translocation in an infertile man: a case report.

J Assist Reprod Genet 2013 Dec 24;30(12):1553-8. Epub 2013 Oct 24.

Institute of Human Genetics, University of Münster, Vesaliusweg 12-14, 48149, Münster, Germany.

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http://dx.doi.org/10.1007/s10815-013-0122-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3843169PMC
December 2013

Derivation and expansion using only small molecules of human neural progenitors for neurodegenerative disease modeling.

PLoS One 2013 22;8(3):e59252. Epub 2013 Mar 22.

Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, North Rhine Westphalia, Germany.

Phenotypic drug discovery requires billions of cells for high-throughput screening (HTS) campaigns. Because up to several million different small molecules will be tested in a single HTS campaign, even small variability within the cell populations for screening could easily invalidate an entire campaign. Neurodegenerative assays are particularly challenging because neurons are post-mitotic and cannot be expanded for implementation in HTS. Therefore, HTS for neuroprotective compounds requires a cell type that is robustly expandable and able to differentiate into all of the neuronal subtypes involved in disease pathogenesis. Here, we report the derivation and propagation using only small molecules of human neural progenitor cells (small molecule neural precursor cells; smNPCs). smNPCs are robust, exhibit immortal expansion, and do not require cumbersome manual culture and selection steps. We demonstrate that smNPCs have the potential to clonally and efficiently differentiate into neural tube lineages, including motor neurons (MNs) and midbrain dopaminergic neurons (mDANs) as well as neural crest lineages, including peripheral neurons and mesenchymal cells. These properties are so far only matched by pluripotent stem cells. Finally, to demonstrate the usefulness of smNPCs we show that mDANs differentiated from smNPCs with LRRK2 G2019S are more susceptible to apoptosis in the presence of oxidative stress compared to wild-type. Therefore, smNPCs are a powerful biological tool with properties that are optimal for large-scale disease modeling, phenotypic screening, and studies of early human development.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0059252PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3606479PMC
September 2013

Comprehensive sequence analysis of the NR5A1 gene encoding steroidogenic factor 1 in a large group of infertile males.

Eur J Hum Genet 2013 Sep 9;21(9):1012-5. Epub 2013 Jan 9.

Institute of Human Genetics, University of Münster, Münster, Germany.

The steroidogenic factor 1 (SF1) protein, encoded by the NR5A1 gene, plays a central role in gonadal development and steroidogenesis. Mutations in NR5A1 were first described in patients with primary adrenal insufficiency and 46,XY disorders of sexual development and later also in men with hypospadias, bilateral anorchia and micropenis and women with primary ovarian insufficiency. Recently, heterozygous missense mutations were found in 4% of infertile men with unexplained reduced sperm counts living in France, but all mutation carriers were of non-Caucasian ancestry. Therefore, we performed a comprehensive NR5A1 sequence analysis in 488 well-characterised predominantly Caucasian patients with azoo- or severe oligozoospermia. Two-hundred-thirty-seven men with normal semen parameters were sequenced as controls. In addition to several synonymous variants of unclear pathogenicity, three heterozygous missense mutations predicted to be damaging to SF1 protein function were identified. The andrological phenotype in infertile but otherwise healthy mutation carriers seems variable. In conclusion, mutations altering SF1 protein function and causing spermatogenic failure are also found in men of German origin, but the prevalence seems markedly lower than in other populations.
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http://dx.doi.org/10.1038/ejhg.2012.290DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3746266PMC
September 2013