Hackett TA
PMID: 30315630 | DOI: 10.1002/ar.23907
In the brain, purines such as ATP and adenosine can function as neurotransmitters and co-transmitters, or serve as signals in neuron-glial interactions. In thalamocortical (TC) projections to sensory cortex, adenosine functions as a negative regulator of glutamate release via activation of the presynaptic adenosine A1 receptor (A1 R). In the auditory forebrain, restriction of A1 R-adenosine signaling in medial geniculate (MG) neurons is sufficient to extend LTP, LTD, and tonotopic map plasticity in adult mice for months beyond the critical period. Interfering with adenosine signaling in primary auditory cortex (A1) does not contribute to these forms of plasticity, suggesting regional differences in the roles of A1 R-mediated adenosine signaling in the forebrain. To advance understanding of the circuitry, in situ hybridization was used to localize neuronal and glial cell types in the auditory forebrain that express A1 R transcripts (Adora1), based on co-expression with cell-specific markers for neuronal and glial subtypes. In A1, Adora1 transcripts were concentrated in L3/4 and L6 of glutamatergic neurons. Subpopulations of GABAergic neurons, astrocytes, oligodendrocytes, and microglia expressed lower levels of Adora1. In MG, Adora1 was expressed by glutamatergic neurons in all divisions, and subpopulations of all glial classes. The collective findings imply that A1 R-mediated signaling broadly extends to all subdivisions of auditory cortex and MG. Selective expression by neuronal and glial subpopulations suggests that experimental manipulations of A1 R-adenosine signaling could impact several cell types, depending on their location. Strategies to target Adora1 in specific cell types can be developed from the data generated here.
Feigin, CY;Moreno, JA;Ramos, R;Mereby, SA;Alivisatos, A;Wang, W;van Amerongen, R;Camacho, J;Rasweiler, JJ;Behringer, RR;Ostrow, B;Plikus, MV;Mallarino, R;
PMID: 36961889 | DOI: 10.1126/sciadv.ade7511
Lateral flight membranes, or patagia, have evolved repeatedly in diverse mammalian lineages. While little is known about patagium development, its recurrent evolution may suggest a shared molecular basis. By combining transcriptomics, developmental experiments, and mouse transgenics, we demonstrate that lateral Wnt5a expression in the marsupial sugar glider (Petaurus breviceps) promotes the differentiation of its patagium primordium. We further show that this function of Wnt5a reprises ancestral roles in skin morphogenesis predating mammalian flight and has been convergently used during patagium evolution in eutherian bats. Moreover, we find that many genes involved in limb development have been redeployed during patagium outgrowth in both the sugar glider and bat. Together, our findings reveal that deeply conserved genetic toolkits contribute to the evolutionary transition to flight in mammals.
DETERMINATION OF SINGLE NUCLEOTIDE POLYMORPHISM (RS566926) OF WNT5A IN NONSYNDROMIC CLEFT LIP AND PALATE IN A PAKISTANI POPULATION
Oral Surgery, Oral Medicine, Oral Pathology and Oral Radiology
Anjum, R;Mehmood, S;Nagi, A;Shahzad, M;Chuadhry, S;
| DOI: 10.1016/j.oooo.2021.03.042
Background Orofacial clefts are the most common birth defects affecting 1 in 750 live births worldwide. Various genetic loci to be involved in nonsyndromic cleft lip and palate has been identified with a variation among populations. Wnt5a is expressed in the frontonasal prominence and maxillary process, which fuse to form the primary palate. Therefore, its dysregulation can lead to certain birth defects along with other diseases. Single nucleotide polymorphism (rs566926) in Wnt5A shows a significant association with nonsyndromic cleft lip and palate in Brazilian and European American populations. Objective The aim of the present study was to describe single nucleotide polymorphism (SNP; rs566926) in patients with nonsyndromic cleft lip and palate in a Pakistani population. Methods This study was conducted on 120 patients with nonsyndromic cleft lip and palate. Demographics and phenotypes were noted. Blood samples were collected in ethylenediaminetetraacetic acid vials. DNA was extracted followed by conventional polymerase chain reaction. SNP (566926) was determined by Sanger sequencing. Data were analyzed using NCBI Blast and SPSS (24.0). Results The mean age of n = 30 patients was 51.33 ± 61.33 months. Sixty percent were male and 40% were female. Regarding cleft types, 70% were both cleft lip and palate, 26% cleft lip only, and 3.3% cleft palate only. Heterozygous polymorphism (T/G) was seen in 33.3% of patients with both cleft lip and palate with bilateral involvement and heterozygous polymorphism (T) was seen in 16.6%. Conclusions SNP in the WNT5A gene is associated with cleft lip and palate, supporting its involvement in pathogenesis of cleft lip and palate. Further studies are recommended to determine the role of Wnt5a genes during craniofacial development.
Mizutani M, Wu JC, Nusse R.
PMID: - | DOI: 10.1161/JAHA.115.002457
Background The adult mammalian heart responds to cardiac injury by formation of persistent fibrotic scar that eventually leads to heart failure. In contrast, the neonatal mammalian heart reacts to injury by the development of transient fibrotic tissue that is eventually replaced by regenerated cardiomyocytes. How fibrosis occurs in the neonatal mammalian heart remains unknown. To start elucidating the molecular underpinnings of neonatal cardiac fibrosis, we investigated Wnt signaling in the neonatal heart after cryoinjury.
Methods and Results Using expression of the Wnt target gene Axin2 as an indicator of Wnt/β‐catenin signaling activation, we discovered that epicardial cells in the ventricles are responsive to Wnt in the uninjured neonatal heart. Lineage‐tracing studies of these Wnt‐responsive epicardial cells showed that they undergo epithelial‐to‐mesenchymal transition and infiltrate into the subepicardial space and exhibit fibroblast phenotypes after injury. In addition, we showed that—similar to adult ischemic injury—neonatal cryoinjury results in activation of Wnt signaling in cardiac fibroblasts near injured areas. Furthermore, through in situ hybridization of all 19 Wnt ligands in injured neonatal hearts, we observed upregulation of Wnt ligands (Wnt2b, Wnt5a, and Wnt9a) that had not been implicated in the adult cardiac injury response.
Conclusions These results demonstrate that cryoinjury in neonatal heart leads to the formation of fibrotic tissue that involves Wnt‐responsive epicardial cells undergoing epithelial‐to‐mesenchymal transition to give rise to fibroblasts and activation of Wnt signaling in resident cardiac fibroblasts.
WNT signaling in pre-granulosa cells is required for ovarian folliculogenesis and female fertility
Development (Cambridge, England)
Habara, O;Logan, CY;Kanai-Azuma, M;Nusse, R;Takase, HM;
PMID: 33914868 | DOI: 10.1242/dev.198846
In mammalian ovaries, immature oocytes are reserved in primordial follicles until their activation for potential ovulation. Precise control of primordial follicle activation (PFA) is essential for reproduction, but how this is achieved is unclear. Here, we show that canonical wingless-type MMTV integration site family (WNT) signaling is pivotal for pre-granulosa cell (pre-GC) activation during PFA. We identified several WNT ligands expressed in pre-GCs that act in an autocrine manner. Inhibition of WNT secretion from pre-GCs/GCs by conditional knockout (cKO) of the wntless (Wls) gene led to female infertility. In Wls cKO mice, GC layer thickness was greatly reduced in growing follicles, which resulted in impaired oocyte growth with both an abnormal, sustained nuclear localization of forkhead box O3 (FOXO3) and reduced phosphorylation of ribosomal protein S6 (RPS6). Constitutive stabilization of β-catenin (CTNNB1) in pre-GCs/GCs induced morphological changes of pre-GCs from a squamous into a cuboidal form, though it did not influence oocyte activation. Our results reveal that canonical WNT signaling plays a permissive role in the transition of pre-GCs to GCs, which is an essential step to support oocyte growth.
Cellular and molecular gastroenterology and hepatology
Kim, TY;Kim, S;Kim, Y;Lee, YS;Lee, S;Lee, SH;Kweon, MN;
PMID: 34971821 | DOI: 10.1016/j.jcmgh.2021.12.015
Dietary signals are known to modulate stemness and tumorigenicity of intestinal progenitors; however, the impact of a high-fat diet (HFD) on the intestinal stem cell (ISC) niche and its association with colorectal cancer remains unclear. Thus, we aimed to investigate how a HFD affects the ISC niche and its regulatory factors.Mice were fed a purified diet (PD) or HFD for 2 months. The expression levels of ISC-related markers, ISC-supportive signals, and Wnt2b were assessed with real-time quantitative polymerase chain reaction, in situ hybridization, and immunofluorescence staining. RNA sequencing and metabolic function were analyzed in mesenchymal stromal cells (MSCs) from PD- and HFD-fed mice. Fecal microbiota were analyzed by 16s rRNA sequencing. Bile salt hydrolase activity and bile acid (BA) levels were measured.We found that expression of CD44 and Wnt signal-related genes was higher in the colonic crypts of HFD-fed mice than in those fed a PD. Within the ISC niche, MSCs were expanded and secreted predominant levels of Wnt2b in the colon of HFD-fed mice. Of note, increased energy metabolism and cancer-associated fibroblast (CAF)-like properties were found in the colonic MSCs of HFD-fed mice. Moreover, colonic MSCs from HFD-fed mice promoted the growth of tumorigenic properties and accelerated the expression of cancer stem cell (CSC)-related markers in colon organoids. In particular, production of primary and secondary BAs was increased through the expansion of bile salt hydrolase-encoding bacteria in HFD-fed mice. Most importantly, BAs-FXR interaction stimulated Wnt2b production in colonic CAF-like MSCs.HFD-induced colonic CAF-like MSCs play an indispensable role in balancing the properties of CSCs through activation of the BAs-FXR axis.
Rehman, R;Miller, M;Krishnamurthy, SS;Kjell, J;Elsayed, L;Hauck, SM;Olde Heuvel, F;Conquest, A;Chandrasekar, A;Ludolph, A;Boeckers, T;Mulaw, MA;Goetz, M;Morganti-Kossmann, MC;Takeoka, A;Roselli, F;
PMID: 36577378 | DOI: 10.1016/j.celrep.2022.111867
The complexity of signaling events and cellular responses unfolding in neuronal, glial, and immune cells upon traumatic brain injury (TBI) constitutes an obstacle in elucidating pathophysiological links and targets for intervention. We use array phosphoproteomics in a murine mild blunt TBI to reconstruct the temporal dynamics of tyrosine-kinase signaling in TBI and then scrutinize the large-scale effects of perturbation of Met/HGFR, VEGFR1, and Btk signaling by small molecules. We show Met/HGFR as a selective modifier of early microglial response and that Met/HGFR blockade prevents the induction of microglial inflammatory mediators, of reactive microglia morphology, and TBI-associated responses in neurons and vasculature. Both acute and prolonged Met/HGFR inhibition ameliorate neuronal survival and motor recovery. Early elevation of HGF itself in the cerebrospinal fluid of TBI patients suggests that this mechanism has translational value in human subjects. Our findings identify Met/HGFR as a modulator of early neuroinflammation in TBI with promising translational potential.
Trobisch, T;Zulji, A;Stevens, NA;Schwarz, S;Wischnewski, S;Öztürk, M;Perales-Patón, J;Haeussler, M;Saez-Rodriguez, J;Velmeshev, D;Schirmer, L;
PMID: 36112223 | DOI: 10.1007/s00401-022-02497-2
Multiple sclerosis (MS) is a multifocal and progressive inflammatory disease of the central nervous system (CNS). However, the compartmentalized pathology of the disease affecting various anatomical regions including gray and white matter and lack of appropriate disease models impede understanding of the disease. Utilizing single-nucleus RNA-sequencing and multiplex spatial RNA mapping, we generated an integrated transcriptomic map comprising leukocortical, cerebellar and spinal cord areas in normal and MS tissues that captures regional subtype diversity of various cell types with an emphasis on astrocytes and oligodendrocytes. While we found strong cross-regional diversity among glial subtypes in control tissue, regional signatures become more obscure in MS. This suggests that patterns of transcriptomic changes in MS are shared across regions and converge on specific pathways, especially those regulating cellular stress and immune activation. In addition, we found evidence that a subtype of white matter oligodendrocytes appearing across all three CNS regions adopt pro-remyelinating gene signatures in MS. In summary, our data suggest that cross-regional transcriptomic glial signatures overlap in MS, with different reactive glial cell types capable of either exacerbating or ameliorating pathology.
Snowball J, Ambalavanan M, Whitsett J, Sinner D.
PMID: 26093309 | DOI: 10.1016/j.ydbio.2015.06.009.
Tracheobronchomalacia is a common congenital defect in which the walls of the trachea and bronchi lack of adequate cartilage required for support of the airways. Deletion of Wls, a cargo receptor mediating Wnt ligand secretion, in the embryonic endoderm using ShhCre mice inhibited formation of tracheal-bronchial cartilaginous rings. The normal dorsal-ventral patterning of tracheal mesenchyme was lost. Smooth muscle cells, identified by Acta2 staining, were aberrantly located in ventral mesenchyme of the trachea, normally the region of Sox9 expression in cartilage progenitors. Wnt/β-catenin activity, indicated by Axin2 LacZ reporter, was decreased in tracheal mesenchyme of Wlsf/f;ShhCre/+ embryos. Proliferation of chondroblasts was decreased and reciprocally, proliferation of smooth muscle cells was increased in Wlsf/f;ShhCre/+ tracheal tissue. Expression of Tbx4, Tbx5, Msx1 and Msx2, known to mediate cartilage and muscle patterning, were decreased in tracheal mesenchyme of Wlsf/f;ShhCre/+ embryos. Ex vivo studies demonstrated that Wnt7b and Wnt5a, expressed by the epithelium of developing trachea, and active Wnt/β-catenin signaling are required for tracheal chondrogenesis before formation of mesenchymal condensations. In conclusion, Wnt ligands produced by the tracheal epithelium pattern the tracheal mesenchyme via modulation of gene expression and cell proliferation required for proper tracheal cartilage and smooth muscle differentiation.
Ma, S;Skarica, M;Li, Q;Xu, C;Risgaard, RD;Tebbenkamp, ATN;Mato-Blanco, X;Kovner, R;Krsnik, Ž;de Martin, X;Luria, V;Martí-Pérez, X;Liang, D;Karger, A;Schmidt, DK;Gomez-Sanchez, Z;Qi, C;Gobeske, KT;Pochareddy, S;Debnath, A;Hottman, CJ;Spurrier, J;Teo, L;Boghdadi, AG;Homman-Ludiye, J;Ely, JJ;Daadi, EW;Mi, D;Daadi, M;Marín, O;Hof, PR;Rasin, MR;Bourne, J;Sherwood, CC;Santpere, G;Girgenti, MJ;Strittmatter, SM;Sousa, AMM;Sestan, N;
PMID: 36007006 | DOI: 10.1126/science.abo7257
The granular dorsolateral prefrontal cortex (dlPFC) is an evolutionary specialization of primates that is centrally involved in cognition. Here, we assessed over 600,000 single-nucleus transcriptomes from adult human, chimpanzee, macaque, and marmoset dlPFC. While most transcriptomically-defined cell subtypes are conserved, we detected several only in some species and substantial species-specific molecular differences across homologous neuronal, glial and non-neural subtypes. The latter are exemplified by human-specific switching between expression of the neuropeptide somatostatin (SST) and tyrosine hydroxylase (TH), the rate-limiting enzyme in dopamine production, in certain interneurons, and also by expression of the neuropsychiatric risk gene FOXP2, which is human-specific in microglia and primate-specific in layer-4 granular neurons. We generated a comprehensive survey of dlPFC cellular repertoire and its shared and divergent features in anthropoid primates.
Guerrero-Juarez, CF;Lee, GH;Liu, Y;Wang, S;Karikomi, M;Sha, Y;Chow, RY;Nguyen, TTL;Iglesias, VS;Aasi, S;Drummond, ML;Nie, Q;Sarin, K;Atwood, SX;
PMID: 35687691 | DOI: 10.1126/sciadv.abm7981
How basal cell carcinoma (BCC) interacts with its tumor microenvironment to promote growth is unclear. We use singe-cell RNA sequencing to define the human BCC ecosystem and discriminate between normal and malignant epithelial cells. We identify spatial biomarkers of tumors and their surrounding stroma that reinforce the heterogeneity of each tissue type. Combining pseudotime, RNA velocity-PAGA, cellular entropy, and regulon analysis in stromal cells reveals a cancer-specific rewiring of fibroblasts, where STAT1, TGF-β, and inflammatory signals induce a noncanonical WNT5A program that maintains the stromal inflammatory state. Cell-cell communication modeling suggests that tumors respond to the sudden burst of fibroblast-specific inflammatory signaling pathways by producing heat shock proteins, whose expression we validated in situ. Last, dose-dependent treatment with an HSP70 inhibitor suppresses in vitro vismodegib-resistant BCC cell growth, Hedgehog signaling, and in vivo tumor growth in a BCC mouse model, validating HSP70's essential role in tumor growth and reinforcing the critical nature of tumor microenvironment cross-talk in BCC progression.
Liu, Y;Guerrero-Juarez, C;Xiao, F;Shettigar, N;Ramos, R;Kuan, C;Lin, Y;de Jesus Martinez Lomeli, L;Park, J;Oh, J;Liu, R;Lin, S;Tartaglia, M;Yang, R;Yu, Z;Nie, Q;Li, J;Plikus, M;
| DOI: 10.1016/j.devcel.2022.06.005
Hair follicle stem cells are regulated by dermal papilla fibroblasts, their principal signaling niche. Overactivation of Hedgehog signaling in the niche dramatically accelerates hair growth and induces follicle multiplication in mice. On single-cell RNA sequencing, dermal papilla fibroblasts increase heterogeneity to include new Wnt5ahigh states. Transcriptionally, mutant fibroblasts activate regulatory networks for Gli1, Alx3, Ebf1, Hoxc8, Sox18, and Zfp239. These networks jointly upregulate secreted factors for multiple hair morphogenesis and hair-growth-related pathways. Among these is non-conventional TGF-β ligand Scube3. We show that in normal mouse skin, Scube3 is expressed only in dermal papillae of growing, but not in resting follicles. SCUBE3 protein microinjection is sufficient to induce new hair growth, and pharmacological TGF-β inhibition rescues mutant hair hyper-activation phenotype. Moreover, dermal-papilla-enriched expression of SCUBE3 and its growth-activating effect are partially conserved in human scalp hair follicles. Thus, Hedgehog regulates mesenchymal niche function in the hair follicle via SCUBE3/TGF-β mechanism.
