Transcription Factor GATA6: A Novel Marker and Putative Inducer of Ductal Metaplasia in Biliary Atresia.

Am J Physiol Gastrointest Liver Physiol.

2018 Feb 01

Soini T, Pihlajoki M, Andersson N, Lohi J, Huppert KA, Rudnick DA, Huppert SS, Wilson DB, Pakarinen MP, Heikinheimo M.
PMID: 29388792 | DOI: 10.1152/ajpgi.00362.2017

Biliary atresia (BA), a neonatal liver disease, is characterized by obstruction of extrahepatic bile ducts with subsequent cholestasis, inflammation, and progressive liver fibrosis. To gain insights into the pathophysiology of BA, we focused attention on GATA6, a transcription factor implicated in biliary development. Early in fetal development GATA6 expression is evident in cholangiocytes and hepatocytes, but by late gestation it is extinguished in hepatocytes. Utilizing a unique set of BA liver samples collected before and after successful portoenterostomy (PE), we found that GATA6 expression is markedly upregulated in hepatocytes of patients with BA compared to healthy and cholestatic disease controls. This upregulation is recapitulated in two murine models simulating bile duct obstruction and intrahepatic bile ductule expansion. GATA6 expression in BA livers correlates with two established negative prognostic indicators (age at PE, degree of intrahepatic bile ductule expansion) and decreases after normalization of serum bilirubin by PE. GATA6 expression in BA livers correlates with expression of known regulators of cholangiocyte differentiation ( JAGGED1, HNF1β, and HNF6). These same genes are upregulated after enforced expression of GATA6 in human hepatocyte cell models. In conclusion, GATA6 is a novel marker and a putative driver of hepatocyte-cholangiocyte metaplasia in BA and its expression in hepatocytes is downregulated after successful PE.

A distinct regulatory region of the Bmp5 locus activates gene expression following adult bone fracture or soft tissue injury

Bone. 2015 Apr 14.

Guenther CA, Wang Z, Li E, Tran MC, Logan CY, Nusse R, Pantalena-Filho L, Yang GP, Kingsley DM.
PMID: 25886903 | DOI: 10.1016/j.bone.2015.04.010.

Bone morphogenetic proteins (BMPs) are key signaling molecules required for normal development of bones and other tissues. Previous studies have shown that null mutations in the mouse Bmp5 gene alter the size, shape and number of multiple bone and cartilage structures during development. Bmp5 mutations also delay healing of rib fractures in adult mutants, suggesting that the same signals used to pattern embryonic bone and cartilage are also reused during skeletal regeneration and repair. Despite intense interest in BMPs as agents for stimulating bone formation in clinical applications, little is known about the regulatory elements that control developmental or injury-induced BMP expression. To compare the DNA sequences that activate gene expression during embryonic bone formation and following acute injuries in adult animals, we assayed regions surrounding the Bmp5 gene for their ability to stimulate lacZ reporter gene expression in transgenic mice. Multiple genomic fragments, distributed across the Bmp5 locus, collectively coordinate expression in discrete anatomic domains during normal development, including in embryonic ribs. In contrast, a distinct regulatory region activated expression following rib fracture in adult animals. The same injury control region triggered gene expression in mesenchymal cells following tibia fracture, in migrating keratinocytes following dorsal skin wounding, and in regenerating epithelial cells following lung injury. The Bmp5 gene thus contains an "injury response" control region that is distinct from embryonic enhancers, and that is activated by multiple types of injury in adult animals.

Mesothelium-Derived Factors Shape GATA6-Positive Large Cavity Macrophages

Journal of immunology (Baltimore, Md. : 1950)

2022 Jul 22

Lai, CW;Bagadia, P;Barisas, DAG;Jarjour, NN;Wong, R;Ohara, T;Muegge, BD;Lu, Q;Xiong, S;Edelson, BT;Murphy, KM;Stappenbeck, TS;
PMID: 35868637 | DOI: 10.4049/jimmunol.2200278

The local microenvironment shapes macrophage differentiation in each tissue. We hypothesized that in the peritoneum, local factors in addition to retinoic acid can support GATA6-driven differentiation and function of peritoneal large cavity macrophages (LCMs). We found that soluble proteins produced by mesothelial cells lining the peritoneal cavity maintained GATA6 expression in cultured LCMs. Analysis of global gene expression of isolated mesothelial cells highlighted mesothelin (Msln) and its binding partner mucin 16 (Muc16) as candidate secreted ligands that potentially regulate GATA6 expression in peritoneal LCMs. Mice deficient for either of these molecules showed diminished GATA6 expression in peritoneal and pleural LCMs that was most prominent in aged mice. The more robust phenotype in older mice suggested that monocyte-derived macrophages were the target of Msln and Muc16. Cell transfer and bone marrow chimera experiments supported this hypothesis. We found that lethally irradiated Msln-/- and Muc16-/- mice reconstituted with wild-type bone marrow had lower levels of GATA6 expression in peritoneal and pleural LCMs. Similarly, during the resolution of zymosan-induced inflammation, repopulated peritoneal LCMs lacking expression of Msln or Muc16 expressed diminished GATA6. These data support a role for mesothelial cell-produced Msln and Muc16 in local macrophage differentiation within large cavity spaces such as the peritoneum. The effect appears to be most prominent on monocyte-derived macrophages that enter into this location as the host ages and also in response to infection.

Single-cell RNA sequencing of human nail unit defines RSPO4 onychofibroblasts and SPINK6 nail epithelium

Communications biology

2021 Jun 07

Kim, HJ;Shim, JH;Park, JH;Shin, HT;Shim, JS;Jang, KT;Park, WY;Lee, KH;Kwon, EJ;Jang, HS;Yang, H;Lee, JH;Yang, JM;Lee, D;
PMID: 34099859 | DOI: 10.1038/s42003-021-02223-w

Research on human nail tissue has been limited by the restricted access to fresh specimen. Here, we studied transcriptome profiles of human nail units using polydactyly specimens. Single-cell RNAseq with 11,541 cells from 4 extra digits revealed nail-specific mesenchymal and epithelial cell populations, characterized by RSPO4 (major gene in congenital anonychia) and SPINK6, respectively. In situ RNA hybridization demonstrated the localization of RSPO4, MSX1 and WIF1 in onychofibroblasts suggesting the activation of WNT signaling. BMP-5 was also expressed in onychofibroblasts implicating the contribution of BMP signaling. SPINK6 expression distinguished the nail-specific keratinocytes from epidermal keratinocytes. RSPO4+ onychofibroblasts were distributed at close proximity with LGR6+ nail matrix, leading to WNT/β-catenin activation. In addition, we demonstrated RSPO4 was overexpressed in the fibroblasts of onychomatricoma and LGR6 was highly expressed at the basal layer of the overlying epithelial component, suggesting that onychofibroblasts may play an important role in the pathogenesis of onychomatricoma.

GATA6 and CK5 stratify the survival of patients with pancreatic cancer undergoing neoadjuvant chemotherapy

Modern Pathology

2023 Jan 01

Kokumai, T;Omori, Y;Ishida, M;Ohtsuka, H;Mizuma, M;Nakagawa, K;Chiho, M;Ono, Y;Mizukami, Y;Miura, S;Kume, K;Masamune, A;Morikawa, T;Unno, M;Furukawa, T;
| DOI: 10.1016/j.modpat.2023.100102

Relevant protein expression of GATA6, CK5, vimentin, and mucins using immunohistochemistry was assessed for predicting the prognosis and chemotherapy efficacy in pancreatic cancer (PC). The protein expression was examined in 159 PCs resected after neoadjuvant chemotherapy (NAC-PCs) with 120 matched biopsy specimens taken before NAC. KRAS mutations were assessed by digital PCR. NAC-PCs were classified by GATA6 expression initially and CK5 expression subsequently into four types, i.e., classical type (n = 22) showing GATA6-high (≧ 50%)/CK5-low (< 10%), hybrid type (n = 45) showing GATA6-high/CK5-high (≧ 10%), basal-like type (n = 53) showing GATA6-low (< 50%)/CK5-high (≧ 30%), and null type (n = 39) showing GATA6-low/CK5-low (< 30%), which resulted in a well-stratification of the patients’ prognosis. The classical type showed the most favorable prognosis, while the null type showed the worst prognosis (multivariate hazard ratio 3.56, 95% confidence interval (CI) 1.63−7.77, p = 0.0015). The hybrid and basal-like types were in between. The risk for hepatic recurrence was lower in the classical type than null (multivariate odds ratio (mOR) 0.18, CI 0.04−0.96, p = 0.0449) and basal-like (mOR 0.24, CI 0.05−1.16, p = 0.0750) types. In contrast, the risk for loco-regional recurrence was higher in classical type than the basal-like type (mOR 5.03, CI 1.20−21.1, p = 0.0272). The hybrid type was subclassified into transition and co-expression patterns with different gastric mucin expression. Vimentin-high (≧ 10%, n = 30) in pre-NAC-PC tissues was associated with poor prognosis (p = 0.0256). Phenotypic transitions between pre- and post-NAC were common (73/120; 61%). PCs with NAC regression grades 2 and 3 showed a transition to poorer prognostic phenotypes (p = 0.0497). KRAS mutations were not associated with these phenotypes. In conclusion, GATA6 and CK5 immunohistochemical expression phenotypes may stratify the survival of NAC-PCs and reflect post-NAC phenotypic transitions associated with poor prognosis. Prompt evaluation of immunohistochemical phenotypes may contribute to designing a precision therapeutic strategy for PC patients.

Structural Remodeling of the Human Colonic Mesenchyme in Inflammatory Bowel Disease

Cell

2018 Sep 27

Kinchen J, Chen HH, Parikh K, Antanaviciute A, Jagielowicz M, Fawkner-Corbett D, Ashley N, Cubitt L, Mellado-Gomez E, Attar M, Sharma E, Wills Q, Bowden R, Richter FC, Ahern D, Puri KD, Henault J, Gervais F, Koohy H, Simmons A.
PMID: - | DOI: 10.1016/j.cell.2018.08.067

Intestinal mesenchymal cells play essential roles in epithelial homeostasis, matrix remodeling, immunity, and inflammation. But the extent of heterogeneity within the colonic mesenchyme in these processes remains unknown. Using unbiased single-cell profiling of over 16,500 colonic mesenchymal cells, we reveal four subsets of fibroblasts expressing divergent transcriptional regulators and functional pathways, in addition to pericytes and myofibroblasts. We identified a niche population located in proximity to epithelial crypts expressing SOX6, F3 (CD142), and WNT genes essential for colonic epithelial stem cellfunction. In colitis, we observed dysregulation of this niche and emergence of an activated mesenchymal population. This subset expressed TNF superfamily member 14 (TNFSF14), fibroblastic reticular cell-associated genes, IL-33, and Lysyl oxidases. Further, it induced factors that impaired epithelial proliferation and maturation and contributed to oxidative stress and disease severity in vivo. Our work defines how the colonic mesenchyme remodels to fuel inflammation and barrier dysfunction in IBD.

Genomics-Driven Precision Medicine for Advanced Pancreatic Cancer: Early Results from the COMPASS Trial

Clin Cancer Res.

2017 Dec 29

Aung KL, Fischer SE, Denroche RE, Jang GH, Dodd A, Creighton S, Southwood B, Liang SB, Chadwick D, Zhang A, O'Kane GM, Albaba H, Moura S, Grant RC, Miller JK, Mbabaali F, Pasternack D, Lungu IM, Bartlett JMS, Ghai S, Lemire M, Holter S, Connor AA, Moffitt
PMID: 29288237 | DOI: 10.1158/1078-0432.CCR-17-2994

Abstract

Purpose: To perform real-time whole genome sequencing (WGS) and RNA sequencing (RNASeq) of advanced pancreatic ductal adenocarcinoma (PDAC) to identify predictive mutational and transcriptional features for better treatment selection.Experimental Design:Patients with advanced PDAC were prospectively recruited prior to first-line combination chemotherapy. Fresh tumor tissue was acquired by image-guided percutaneous core biopsy for WGS and RNASeq. Laser capture microdissection was performed for all cases. Primary endpoint was feasibility to report WGS results prior to first disease assessment CT scan at 8 weeks. The main secondary endpoint was discovery of patient subsets with predictive mutational and transcriptional signatures.Results: Sixty-three patients underwent a tumor biopsy between December 2015 and June 2017. WGS and RNASeq were successful in 62 (98%) and 60 (95%), respectively. Genomic results were reported at a median of 35 days (range, 19-52 days) from biopsy, meeting the primary feasibility endpoint. Objective responses to first-line chemotherapy were significantly better in patients with the classical PDAC RNA subtype compared with those with the basal-like subtype (P = 0.004). The best progression-free survival was observed in those with classical subtype treated with m-FOLFIRINOX. GATA6 expression in tumor measured by RNA in situ hybridization was found to be a robust surrogate biomarker for differentiating classical and basal-like PDAC subtypes. Potentially actionable genetic alterations were found in 30% of patients.Conclusions: Prospective genomic profiling of advanced PDAC is feasible, and our early data indicate that chemotherapy response differs among patients with different genomic/transcriptomic subtypes.

Smooth muscle contributes to the development and function of a layered intestinal stem cell niche

Developmental cell

2023 Mar 08

McCarthy, N;Tie, G;Madha, S;He, R;Kraiczy, J;Maglieri, A;Shivdasani, RA;
PMID: 36924771 | DOI: 10.1016/j.devcel.2023.02.012

Wnt and Rspondin (RSPO) signaling drives proliferation, and bone morphogenetic protein inhibitors (BMPi) impede differentiation, of intestinal stem cells (ISCs). Here, we identify the mouse ISC niche as a complex, multi-layered structure that encompasses distinct mesenchymal and smooth muscle populations. In young and adult mice, diverse sub-cryptal cells provide redundant ISC-supportive factors; few of these are restricted to single cell types. Niche functions refine during postnatal crypt morphogenesis, in part to oppose the dense aggregation of differentiation-promoting BMP+ sub-epithelial myofibroblasts at crypt-villus junctions. Muscularis mucosae, a specialized muscle layer, first appears during this period and supplements neighboring RSPO and BMPi sources. Components of this developing niche are conserved in human fetuses. The in vivo ablation of mouse postnatal smooth muscle increases BMP signaling activity, potently limiting a pre-weaning burst of crypt fission. Thus, distinct and progressively specialized mesenchymal cells together create the milieu that is required to propagate crypts during rapid organ growth and to sustain adult ISCs.

Epicardium-derived cells organize through tight junctions to replenish cardiac muscle in salamanders

Nature cell biology

2022 May 01

Eroglu, E;Yen, CYT;Tsoi, YL;Witman, N;Elewa, A;Joven Araus, A;Wang, H;Szattler, T;Umeano, CH;Sohlmér, J;Goedel, A;Simon, A;Chien, KR;
PMID: 35550612 | DOI: 10.1038/s41556-022-00902-2

The contribution of the epicardium, the outermost layer of the heart, to cardiac regeneration has remained controversial due to a lack of suitable analytical tools. By combining genetic marker-independent lineage-tracing strategies with transcriptional profiling and loss-of-function methods, we report here that the epicardium of the highly regenerative salamander species Pleurodeles waltl has an intrinsic capacity to differentiate into cardiomyocytes. Following cryoinjury, CLDN6+ epicardium-derived cells appear at the lesion site, organize into honeycomb-like structures connected via focal tight junctions and undergo transcriptional reprogramming that results in concomitant differentiation into de novo cardiomyocytes. Ablation of CLDN6+ differentiation intermediates as well as disruption of their tight junctions impairs cardiac regeneration. Salamanders constitute the evolutionarily closest species to mammals with an extensive ability to regenerate heart muscle and our results highlight the epicardium and tight junctions as key targets in efforts to promote cardiac regeneration.

Nail-associated mesenchymal cells contribute to and are essential for dorsal digit tip regeneration

Cell reports

2022 Dec 20

Mahmud, N;Eisner, C;Purushothaman, S;Storer, MA;Kaplan, DR;Miller, FD;
PMID: 36543145 | DOI: 10.1016/j.celrep.2022.111853

Here, we ask why the nail base is essential for mammalian digit tip regeneration, focusing on the inductive nail mesenchyme. We identify a transcriptional signature for these cells that includes Lmx1b and show that the Lmx1b-expressing nail mesenchyme is essential for blastema formation. We use a combination of Lmx1bCreERT2-based lineage-tracing and single-cell transcriptional analyses to show that the nail mesenchyme contributes cells for two pro-regenerative mechanisms. One group of cells maintains their identity and regenerates the new nail mesenchyme. A second group contributes specifically to the dorsal blastema, loses their nail mesenchyme phenotype, acquires a blastema transcriptional state that is highly similar to blastema cells of other origins, and ultimately contributes to regeneration of the dorsal but not ventral dermis and bone. Thus, the regenerative necessity for an intact nail base is explained, at least in part, by a requirement for the inductive nail mesenchyme.

	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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