Zona glomerulosa derived Klotho does not regulate aldosterone synthesis in young mice

Endocrine Abstracts

2021 May 15

Tang, C;Xie, Y;Scapin, A;Loffing, D;Breault, D;Loffing, J;Beuschlein, F;
| DOI: 10.1530/endoabs.73.aep8

Klotho (Kl), initially identified as an antiaging gene, plays a critical role in the regulation of renal and adrenal dependent fluid homeostasis. A previous study reported that haplodeficiency of Kl in mice resulted in increased aldosterone synthase (CYP11B2) expression, elevated plasma aldosterone and high blood pressure. This phenotype was presumed to result from diminished Kl expression in zona glomerulosa (zG) of the adrenal. To examine whether Kl expressed in zG is indeed a critical regulator of aldosterone synthesis, we generated a tamoxifen-inducible, zG-specific mouse model of KI deficiency by crossing Kl-flox mice with Cyp11b2-CreERT mice (zG-Kl). Tamoxifen-treated Cyp11b2-CreERT animals (zG-Cre) served as controls. Rosa26-mTmG reporter mice were used for Cre-dependent lineage-marking. Two weeks after tamoxifen induction, the specificity of the zG-Cre line was verified using immunofluorescence analysis to show that GFP expression was restricted to the zG. RNAScope _in situ_ hybridization revealed a 65% down-regulation of Kl mRNA expression in zG of zG-Kl mice at 12-weeks of age compared to control mice. Despite this, zG-Kl mice exhibited no difference in adrenal Cyp11b2 expression or plasma aldosterone levels compared to control mice independent of sex. These results suggest that zG-derived Kl _per se_ does not significantly regulate aldosterone synthesis in young adult mice. Further studies are required to investigate the role of adrenal Kl in aldosterone synthesis in aged mice.

The MAL Protein, an Integral Component of Specialized Membranes, in Normal Cells and Cancer

Cells

2021 Apr 30

Rubio-Ramos, A;Labat-de-Hoz, L;Correas, I;Alonso, MA;
PMID: 33946345 | DOI: 10.3390/cells10051065

The MAL gene encodes a 17-kDa protein containing four putative transmembrane segments whose expression is restricted to human T cells, polarized epithelial cells and myelin-forming cells. The MAL protein has two unusual biochemical features. First, it has lipid-like properties that qualify it as a member of the group of proteolipid proteins. Second, it partitions selectively into detergent-insoluble membranes, which are known to be enriched in condensed cell membranes, consistent with MAL being distributed in highly ordered membranes in the cell. Since its original description more than thirty years ago, a large body of evidence has accumulated supporting a role of MAL in specialized membranes in all the cell types in which it is expressed. Here, we review the structure, expression and biochemical characteristics of MAL, and discuss the association of MAL with raft membranes and the function of MAL in polarized epithelial cells, T lymphocytes, and myelin-forming cells. The evidence that MAL is a putative receptor of the epsilon toxin of Clostridium perfringens, the expression of MAL in lymphomas, the hypermethylation of the MAL gene and subsequent loss of MAL expression in carcinomas are also presented. We propose a model of MAL as the organizer of specialized condensed membranes to make them functional, discuss the role of MAL as a tumor suppressor in carcinomas, consider its potential use as a cancer biomarker, and summarize the directions for future research.

The Expression and Function of Human Ribonuclease 4 in the Kidney and Urinary Tract

American journal of physiology. Renal physiology

2021 Apr 05

Bender, KI;Schwartz, LL;Cohen, A;Mosquera Vasquez, C;Murtha, MJ;Eichler, T;Thomas, JP;Jackson, AR;Spencer, JD;
PMID: 33818125 | DOI: 10.1152/ajprenal.00592.2020

Antimicrobial peptides are essential host defense mechanisms that prevent urinary tract infections. Recent studies demonstrate that peptides in the Ribonuclease A Superfamily have antimicrobial activity against uropathogens and protect the urinary tract from uropathogenic Escherichia coli (UPEC). Little is known is about the antibacterial function or expression of Ribonuclease 4 in the human urinary tract. Here, we show that full-length recombinant Ribonuclease 4 peptide and synthetic amino-terminal Ribonuclease 4 peptide fragment have antibacterial activity against UPEC and multi-drug resistant UPEC. RNASE4 transcript expression was detected in human kidney and bladder tissue using quantitative real-time polymerase chain reaction. Immunostaining or in situ hybridization localized Ribonuclease 4 expression to proximal tubules, principal and intercalated cells in the kidney's collecting duct, and the bladder urothelium. Urinary Ribonuclease 4 concentrations were quantified in healthy controls and females with a urinary tract infection history. Compared to controls, urinary RNase 4 concentrations were significantly lower in females with a urinary tract infection history. When Ribonuclease 4 was neutralized in human urine or silenced in vitro using small interfering RNA, urinary UPEC replication or attachment to and invasion of urothelial and kidney medullary cells increased. These data show that Ribonuclease 4 has antibacterial activity against UPEC, is expressed in the human urinary tract, and can contribute to host defense against urinary tract infections.

Habenula GPR139 is associated with fear learning in the zebrafish

Scientific reports

2021 Mar 10

Roy, N;Ogawa, S;Maniam, R;Parhar, I;
PMID: 33692406 | DOI: 10.1038/s41598-021-85002-1

G-protein coupled receptor 139 (GPR139) is an evolutionarily conserved orphan receptor, predominantly expressing in the habenula of vertebrate species. The habenula has recently been implicated in aversive response and its associated learning. Here, we tested the hypothesis that GPR139 signalling in the habenula may play a role in fear learning in the zebrafish. We examined the effect of intraperitoneal injections of a human GPR139-selective agonist (JNJ-63533054) on alarm substance-induced fear learning using conditioned place avoidance paradigm, where an aversive stimulus is paired with one compartment, while its absence is associated with the other compartment of the apparatus. The results indicate that fish treated with 1 µg/g body weight of GPR139 agonist displayed no difference in locomotor activity and alarm substance-induced fear response. However, avoidance to fear-conditioned compartment was diminished, which suggests that the agonist blocks the consolidation of contextual fear memory. On the other hand, fish treated with 0.1 µg/g body weight of GPR139 agonist spent a significantly longer time in the unconditioned neutral compartment as compared to the conditioned (punished and unpunished) compartments. These results suggest that activation of GPR139 signalling in the habenula may be involved in fear learning and the decision-making process in the zebrafish.

Development of Testis Cords and the Formation of Efferent Ducts in Xenopus laevis: Differences and Similarities with Other Vertebrates

Sexual development : genetics, molecular biology, evolution, endocrinology, embryology, and pathology of sex determination and differentiation

2021 Mar 04

Li, Y;Li, J;Cai, M;Qin, Z;
PMID: 33662961 | DOI: 10.1159/000513416

The knowledge of testis development in amphibians relative to amniotes remains limited. Here, we used Xenopus laevis to investigate the process of testis cord development. Morphological observations revealed the presence of segmental gonomeres consisting of medullary knots in male gonads at stages 52-53, with no distinct gonomeres in female gonads. Further observations showed that cell proliferation occurs at specific sites along the anterior-posterior axis of the future testis at stage 50, which contributes to the formation of medullary knots. At stage 53, adjacent gonomeres become close to each other, resulting in fusion; then (pre-)Sertoli cells aggregate and form primitive testis cords, which ultimately become testis cords when germ cells are present inside. The process of testis cord formation in X. laevis appears to be more complex than in amniotes. Strikingly, steroidogenic cells appear earlier than (pre-)Sertoli cells in differentiating testes of X. laevis, which differs from earlier differentiation of (pre-)Sertoli cells in amniotes. Importantly, we found that the mesonephros is connected to the testis gonomere at a specific site at early larval stages and that these connections become efferent ducts after metamorphosis, which challenges the previous concept that the mesonephric side and the gonadal side initially develop in isolation and then connect to each other in amphibians and amniotes.

Histopathological features in fatal COVID-19 acute respiratory distress syndrome

Medicina intensiva

2021 Jan 01

Merdji, H;Mayeur, S;Schenck, M;Oulehri, W;Clere-Jehl, R;Cunat, S;Herbrecht, JE;Janssen-Langenstein, R;Nicolae, A;Helms, J;Meziani, F;Chenard, MP;CRICS TRIGGERSEP Group (Clinical Research in Intensive Care, Sepsis Trial Group for Global Evaluation, Research in Sepsis), ;
PMID: 34059216 | DOI: 10.1016/j.medine.2021.02.005

COVID-19 acute respiratory distress syndrome (ARDS) shares the common histological hallmarks with other forms of ARDS. However, the chronology of the histological lesions has not been well established.To describe the chronological histopathological alterations in the lungs of patients with COVID-19 related ARDS.A prospective cohort study was carried out.Intensive Care Unit of a tertiary hospital.The first 22 consecutive COVID-19 deaths.Lung biopsies and histopathological analyses were performed in deceased patients with COVID-19 related ARDS. Clinical data and patient course were evaluated.The median patient age was 66 [63-74] years; 73% were males. The median duration of mechanical ventilation was 17 [8-24] days. COVID-19 induced pulmonary injury was characterized by an exudative phase in the first week of the disease, followed by a proliferative/organizing phase in the second and third weeks, and finally an end-stage fibrosis phase after the third week. Viral RNA and proteins were detected in pneumocytes and macrophages in a very early stage of the disease, and were no longer detected after the second week.Limited sample size.The chronological evolution of COVID-19 lung histopathological lesions seems to be similar to that seen in other forms of ARDS. In particular, lung lesions consistent with potentially corticosteroid-sensitive lesions are seen.

Heightened activation of embryonic megakaryocytes causes aneurysms in the developing brain of mice lacking podoplanin

Blood

2021 Feb 22

Hoover, CM;Kondo, Y;Shao, B;McDaniel, M;Lee, R;McGee, S;Whiteheart, SW;Bergmeier, W;McEver, RP;Xia, L;
PMID: 33619517 | DOI: 10.1182/blood.2020010310

During early embryonic development in mammals, including humans and mice, megakaryocytes first originate from primitive hematopoiesis in the yolk sac. These embryonic megakaryocytes (eMk) circulate in the vasculature with unclear function. Here we report that podoplanin (PDPN), the ligand of C-type lectin-like receptor (CLEC-2) on megakaryocytes/platelets, is temporarily expressed in neural tissue during midgestation in mice. Loss of PDPN or CLEC-2 resulted in aneurysms and spontaneous hemorrhage specifically in the lower diencephalon during midgestation. Surprisingly, more eMks/platelets had enhanced granule release and localized to lower diencephalon in mutant mouse embryos than wild-type littermates prior to hemorrhage. We found that PDPN counteracted the collagen I-induced secretion of angiopoietin-1 from fetal megakaryocytes, which coincided with enhanced TIE2 activation in aneurysm-like sprouts of PDPN-deficient embryos. Blocking platelet activation prevented the PDPN-deficient embryo from developing vascular defects. Our data reveal a new role for PDPN in regulating eMk function during midgestation.

Whole-body senescent cell clearance alleviates age-related brain inflammation and cognitive impairment in mice

Aging cell

2021 Jan 20

Ogrodnik, M;Evans, SA;Fielder, E;Victorelli, S;Kruger, P;Salmonowicz, H;Weigand, BM;Patel, AD;Pirtskhalava, T;Inman, CL;Johnson, KO;Dickinson, SL;Rocha, A;Schafer, MJ;Zhu, Y;Allison, DB;von Zglinicki, T;LeBrasseur, NK;Tchkonia, T;Neretti, N;Passos, JF;Kirkland, JL;Jurk, D;
PMID: 33470505 | DOI: 10.1111/acel.13296

Cellular senescence is characterized by an irreversible cell cycle arrest and a pro-inflammatory senescence-associated secretory phenotype (SASP), which is a major contributor to aging and age-related diseases. Clearance of senescent cells has been shown to improve brain function in mouse models of neurodegenerative diseases. However, it is still unknown whether senescent cell clearance alleviates cognitive dysfunction during the aging process. To investigate this, we first conducted single-nuclei and single-cell RNA-seq in the hippocampus from young and aged mice. We observed an age-dependent increase in p16Ink4a senescent cells, which was more pronounced in microglia and oligodendrocyte progenitor cells and characterized by a SASP. We then aged INK-ATTAC mice, in which p16Ink4a -positive senescent cells can be genetically eliminated upon treatment with the drug AP20187 and treated them either with AP20187 or with the senolytic cocktail Dasatinib and Quercetin. We observed that both strategies resulted in a decrease in p16Ink4a exclusively in the microglial population, resulting in reduced microglial activation and reduced expression of SASP factors. Importantly, both approaches significantly improved cognitive function in aged mice. Our data provide proof-of-concept for senolytic interventions' being a potential therapeutic avenue for alleviating age-associated cognitive impairment.

Single-cell RNA sequencing reveals intratumoral heterogeneity in primary uveal melanomas and identifies HES6 as a driver of the metastatic disease

Cell death and differentiation

2021 Jan 18

Pandiani, C;Strub, T;Nottet, N;Cheli, Y;Gambi, G;Bille, K;Husser, C;Dalmasso, M;Béranger, G;Lassalle, S;Magnone, V;Pédeutour, F;Irondelle, M;Maschi, C;Nahon-Estève, S;Martel, A;Caujolle, JP;Hofman, P;LeBrigand, K;Davidson, I;Baillif, S;Barbry, P;Ballotti, R;Bertolotto, C;
PMID: 33462406 | DOI: 10.1038/s41418-020-00730-7

Intratumor heterogeneity has been recognized in numerous cancers as a major source of metastatic dissemination. In uveal melanomas, the existence and identity of specific subpopulations, their biological function and their contribution to metastasis remain unknown. Here, in multiscale analyses using single-cell RNA sequencing of six different primary uveal melanomas, we uncover an intratumoral heterogeneity at the genomic and transcriptomic level. We identify distinct transcriptional cell states and diverse tumor-associated populations in a subset of the samples. We also decipher a gene regulatory network underlying an invasive and poor prognosis state driven in part by the transcription factor HES6. HES6 heterogenous expression has been validated by RNAscope assays within primary human uveal melanomas, which further unveils the existence of these cells conveying a dismal prognosis in tumors diagnosed with a favorable outcome using bulk analyses. Depletion of HES6 impairs proliferation, migration and metastatic dissemination in vitro and in vivo using the chick chorioallantoic membrane assay, demonstrating the essential role of HES6 in uveal melanomas. Thus, single-cell analysis offers an unprecedented view of primary uveal melanoma heterogeneity, identifies bona fide biomarkers for metastatic cells in the primary tumor, and reveals targetable modules driving growth and metastasis formation. Significantly, our findings demonstrate that HES6 is a valid target to stop uveal melanoma progression.

A role for orphan nuclear receptor liver receptor homolog-1 (LRH-1, NR5A2) in primordial follicle activation

Scientific reports

2021 Jan 13

Meinsohn, MC;Hughes, CHK;Estienne, A;Saatcioglu, HD;Pépin, D;Duggavathi, R;Murphy, BD;
PMID: 33441767 | DOI: 10.1038/s41598-020-80178-4

Liver receptor homolog-1 (NR5A2) is expressed specifically in granulosa cells of developing ovarian follicles where it regulates the late stages of follicle development and ovulation. To establish its effects earlier in the trajectory of follicular development, NR5A2 was depleted from granulosa cells of murine primordial and primary follicles. Follicle populations were enumerated in neonates at postnatal day 4 (PND4) coinciding with the end of the formation of the primordial follicle pool. The frequency of primordial follicles in PND4 conditional knockout (cKO) ovaries was greater and primary follicles were substantially fewer relative to control (CON) counterparts. Ten-day in vitro culture of PND4 ovaries recapitulated in vivo findings and indicated that CON mice developed primary follicles in the ovarian medulla to a greater extent than did cKO animals. Two subsets of primordial follicles were observed in wildtype ovaries: one that expressed NR5A2 and the second in which the transcript was absent. Neither expressed the mitotic marker. KI-67, indicating their developmental quiescence. RNA sequencing on PND4 demonstrated that loss of NR5A2 induced changes in 432 transcripts, including quiescence markers, inhibitors of follicle activation, and regulators of cellular migration and epithelial-to-mesenchymal transition. These experiments suggest that NR5A2 expression poises primordial follicles for entry into the developing pool.

Short prolactin isoforms are expressed in photoreceptors of canine retinas undergoing retinal degeneration

Scientific reports

2021 Jan 11

Sudharsan, R;Murgiano, L;Tang, HY;Olsen, TW;Chavali, VRM;Aguirre, GD;Beltran, WA;
PMID: 33432105 | DOI: 10.1038/s41598-020-80691-6

Prolactin (PRL) hormone functions as a pleiotropic cytokine with a protective role in the retina. We recently identified by transcriptome profiling that PRL is one of the most highly upregulated mRNAs in the retinas of mutant rcd1 (PDE6B) and xlpra2 (RPGR) dogs at advanced stages of photoreceptor disease. In the present study, we have identified the expression of a short PRL isoform that lacks exon 1 in canine retinas and analyzed the time-course of expression and localization of this isoform in the retinas of these two models. Using laser capture microdissection to isolate RNA from each of the retinal cellular layers, we found by qPCR that this short PRL isoform is expressed in photoreceptors of degenerating retinas. We confirmed by in situ hybridization that its expression is localized to the outer nuclear layer and begins shortly after the onset of disease at the time of peak photoreceptor cell death in both models. PRL protein was also detected only in mutant dog retinas. Our results call for further investigations into the role of this novel PRL isoform in retinal degeneration.

High spatial resolution mapping of the mucosal proteome of the gills of Crassostrea virginica: implication in particle processing

The Journal of experimental biology

2021 Jan 11

Pales Espinosa, E;Allam, B;
PMID: 33431594 | DOI: 10.1242/jeb.233361

In the oyster Crassostrea virginica, the organization of the gill allows bidirectional particle transport where a dorsal gill tract directs particles meant to be ingested while a ventral tract collects particles intended to be rejected as pseudofeces. Previous studies showed that the transport of particles in both tracts is mediated by mucus. Consequently, we hypothesized that the nature and/or the quantity of mucosal proteins present in each tract is likely different. Using endoscopy-aided micro-sampling of mucus from each tract followed by multidimensional protein identification technologies, and in situ hybridization, a high spatial resolution mapping of the oyster gill proteome was generated. Results showed the presence in gill mucus of a wide range of molecules involved in non-self recognition and interactions with microbes. Mucus composition was different between the two tracts, with mucus from the ventral tract shown to be rich in mucin-like proteins, providing an explanation of its high viscosity, while mucus from the dorsal tract was found to be enriched in mannose-binding proteins, known to be involved in food particle binding and selection. Overall, this study generated high-resolution proteomes for C. virginica gill mucus and demonstrated that the contrasting functions of the two pathways present on oyster gills are associated with significant differences in their protein makeup.

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	Description
sense Example: Hs-LAG3-sense	Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
Intron# Example: Mm-Htt-intron2	Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
Pool/Pan Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G)	A mixture of multiple probe sets targeting multiple genes or transcripts
No-XSp Example: Hs-PDGFB-No-XMm	Does not cross detect with the species (Sp)
XSp Example: Rn-Pde9a-XMm	designed to cross detect with the species (Sp)
O# Example: Mm-Islr-O1	Alternative design targeting different regions of the same transcript or isoforms
CDS Example: Hs-SLC31A-CDS	Probe targets the protein-coding sequence only
EnEm	Probe targets exons n and m
En-Em	Probe targets region from exon n to exon m
Retired Nomenclature
tvn Example: Hs-LEPR-tv1	Designed to target transcript variant n
ORF Example: Hs-ACVRL1-ORF	Probe targets open reading frame
UTR Example: Hs-HTT-UTR-C3	Probe targets the untranslated region (non-protein-coding region) only
5UTR Example: Hs-GNRHR-5UTR	Probe targets the 5' untranslated region only
3UTR Example: Rn-Npy1r-3UTR	Probe targets the 3' untranslated region only
Pan Example: Pool	A mixture of multiple probe sets targeting multiple genes or transcripts

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