The Glucocorticoid Receptor NR3C1 in Testicular Peritubular Cells is Developmentally Regulated and Linked to the Smooth Muscle-Like Cellular Phenotype

Welter, Harald; Herrmann, Carola; Dellweg, Nils; Missel, Annika; Thanisch, Christiane; Urbanski, Henryk F.; Köhn, F.-M.; Schwarzer, Julia; Müller‐Taubenberger, Annette; Mayerhofer, Artur

doi:10.3390/jcm9040961

Cited by 21 publications

(34 citation statements)

References 30 publications

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“…PA5-101926) was used at a dilution of 1:500. To stain the tissues for Collagen, affinity-purified, polyclonal rabbit anti-Col1 (R1038; Origene) antibody was used at 1:200 dilution ( Welter et al, 2020 ). Negative controls consisted of omission of the primary antibody, of incubation with rabbit IgG or non-immune serum instead of the primary antiserum ( Fig S7 ).…”

Section: Methodsmentioning

confidence: 99%

MALDI-IMS combined with shotgun proteomics identify and localize new factors in male infertility

et al. 2021

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Spermatogenesis is a complex multi-step process involving intricate interactions between different cell types in the male testis. Disruption of these interactions results in infertility. Combination of shotgun tissue proteomics with MALDI imaging mass spectrometry is markedly potent in revealing topological maps of molecular processes within tissues. Here, we use a combinatorial approach on a characterized mouse model of hormone induced male infertility to uncover misregulated pathways. Comparative testicular proteome of wild-type and mice overexpressing human P450 aromatase (AROM+) with pathologically increased estrogen levels unravels gross dysregulation of spermatogenesis and emergence of pro-inflammatory pathways in AROM+ testis. In situ MS allowed us to localize misregulated proteins/peptides to defined regions within the testis. Results suggest that infertility is associated with substantial loss of proteomic heterogeneity, which define distinct stages of seminiferous tubuli in healthy animals. Importantly, considerable loss of mitochondrial factors, proteins associated with late stages of spermatogenesis and steroidogenic factors characterize AROM+ mice. Thus, the novel proteomic approach pinpoints in unprecedented ways the disruption of normal processes in testis and provides a signature for male infertility.

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Section: Methodsmentioning

confidence: 99%

MALDI-IMS combined with shotgun proteomics identify and localize new factors in male infertility

et al. 2021

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“…This implies that CNN1 expression increases with advanced age. For ACTA2, another contractility protein [65,66], expression was reported upon onset of puberty in nonhuman primates, but other age-depending changes are not well known. By binding actins, CNN1 plays an important role in the cytoskeleton [67].…”

Section: Testis Proteomes Of Aged Individuals Show a Broad Increase Of Actin-binding Proteinsmentioning

confidence: 99%

Age-Related Alterations in the Testicular Proteome of a Non-Human Primate

Stöckl¹,

Schmid²,

Flenkenthaler³

et al. 2021

Cells

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Aging of human testis and associated cellular changes is difficult to assess. Therefore, we used a translational, non-human primate model to get insights into underlying cellular and biochemical processes. Using proteomics and immunohistochemistry, we analyzed testicular tissue of young (age 2 to 3) and old (age 10 to 12) common marmosets (Callithrix Jacchus). Using a mass spectrometry-based proteomics approach, we identified 63,124 peptides, which could be assigned to 5924 proteins. Among them, we found proteins specific for germ cells and somatic cells, such as Leydig and Sertoli cells. Quantitative analysis showed 31 differentially abundant proteins, of which 29 proteins were more abundant in older animals. An increased abundance of anti-proliferative proteins, among them CDKN2A, indicate reduced cell proliferation in old testes. Additionally, an increased abundance of several small leucine rich repeat proteoglycans and other extracellular matrix proteins was observed, which may be related to impaired cell migration and fibrotic events. Furthermore, an increased abundance of proteins with inhibitory roles in smooth muscle cell contraction like CNN1 indicates functional alterations in testicular peritubular cells and may mirror a reduced capacity of these cells to contract in old testes.

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“…First strand cDNA synthesis was carried out using dN12 random primers in a volume of 40 µL as described in [25]. Conventional PCR as well as quantitative real-time PCR (qPCR) were performed as described previously [25,26]. PPIA or L19 gene expression served as the internal control.…”

Section: Reverse Transcription (Rt) Conventional and Quantitative Real-time Pcrmentioning

confidence: 99%

“…Immunoblotting of TCam-2 whole-cell lysates was performed as described [21,26]. Protein samples (15 µg) were separated by 10% SDS-PAGE and transferred to a nitrocellulose membrane (pore size 0.2 µm).…”

Section: Western Blottingmentioning

confidence: 99%

Filamin A Orchestrates Cytoskeletal Structure, Cell Migration and Stem Cell Characteristics in Human Seminoma TCam-2 Cells

Welter

Herrmann

Fröhlich

et al. 2020

Cells

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Filamins are large dimeric F-actin cross-linking proteins, crucial for the mechanosensitive properties of a number of cell types. Due to their interaction with a variety of different proteins, they exert important regulatory functions. However, in the human testis the role of filamins has been insufficiently explored. Immunohistochemical staining of human testis samples identified filamin A (FLNA) in spermatogonia and peritubular myoid cells. Investigation of different testicular tumor samples indicated that seminoma also express FLNA. Moreover, mass spectrometric analyses identified FLNA as one of the most abundant proteins in human seminoma TCam-2 cells. We therefore focused on FLNA in TCam-2 cells, and identified by co-immunoprecipitation LAD1, RUVBL1 and DAZAP1, in addition to several cytoskeletal proteins, as interactors of FLNA. To study the role of FLNA in TCam-2 cells, we generated FLNA-deficient cells using the CRISPR/Cas9 system. Loss of FLNA causes an irregular arrangement of the actin cytoskeleton and mechanical instability, impaired adhesive properties and disturbed migratory behavior. Furthermore, transcriptional activity of typical stem cell factors is increased in the absence of FLNA. In summary, our data suggest that FLNA is crucially involved in balancing stem cell characteristics and invasive properties in human seminoma cells and possibly human testicular germ cells.

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The Glucocorticoid Receptor NR3C1 in Testicular Peritubular Cells is Developmentally Regulated and Linked to the Smooth Muscle-Like Cellular Phenotype

Cited by 21 publications

References 30 publications

MALDI-IMS combined with shotgun proteomics identify and localize new factors in male infertility

MALDI-IMS combined with shotgun proteomics identify and localize new factors in male infertility

Age-Related Alterations in the Testicular Proteome of a Non-Human Primate

Filamin A Orchestrates Cytoskeletal Structure, Cell Migration and Stem Cell Characteristics in Human Seminoma TCam-2 Cells

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