All-trans retinoic acid protects mesenchymal stem cells from immune thrombocytopenia by regulating the complement-IL-1β loop.

Haematologica

2019 Jan 24

Zhu X, Wang Y, Jiang Q, Jiang H, Lu J, Wang Y, Kong Y, Chang Y, Xu L, Peng J, Hou M, Huang X, Zhang X.
PMID: 30679324 | DOI: 10.3324/haematol.2018.204446

Peripheral enhanced complement activation has long been considered as one of the major pathogenesis of immune thrombocytopenia. Impaired bone marrow microenvironment, especially the dysfunction of mesenchymal stem cells, has been observed in patients with immune thrombocytopenia. However, the potential role of the complement system involved in impaired bone marrow microenvironment remains poorly understood. Here, bone marrow samples of patients were divided into the MSC-ITP-C+ and MSC-ITP-C- groups based on the deposition of the complement components on the surfaces of mesenchymal stem cells. Reduced and dysfunctional mesenchymal stem cells, characterized by reduced proliferation capacity, increased apoptosis as well as abnormal secretion of interleukin-1β and C-X-C motif chemokine ligand 12, were observed in the MSC-ITP-C+ group. In vitro treatment with all-trans retinoic acid quantitatively and functionally improved MSC-ITP-C+ by upregulating DNA hypermethylation of the interleukin-1β promoter. In vivo studies showed that all-trans retinoic acid could rescue the impaired mesenchymal stem cells to support the thrombopoietic niche in both patients and the murine model with immune thrombocytopenia. Taken together, these results indicate that deficient mesenchymal stem cells mediated by the complement-IL-1β loop play a role in the pathogenesis of immune thrombocytopenia. All-trans retinoic acid represents a promising therapeutic approach in patients with immune thrombocytopenia by repairing impaired mesenchymal stem cells.

Conditional Cell Reprogramming and Air-Liquid Interface Modeling Life Cycle of Oncogenic Viruses (HPV and EBV) in Epithelial Cells and Virus-Associated Human Carcinomas

Viruses

2023 Jun 17

Rani, AQ;Nurmemet, D;Liffick, J;Khan, A;Mitchell, D;Li, J;Zhao, B;Liu, X;
PMID: 37376685 | DOI: 10.3390/v15061388

Several oncogenic viruses are associated with approximately 20% of human cancers. Experimental models are crucial for studying the pathogenicity and biological aspects of oncogenic viruses and their potential mechanisms in tumorigenesis. Current cell models have considerable limitations such as: their low yield, genetic and epigenetic modification, and reduction in tumor heterogeneity during long propagation. Cancer cell lines are limited and not appropriate for studying the viral life cycle, for example, natural viral life cycles of HPV and EBV, and their persistence and latency in epithelial cells are poorly understood, since these processes are highly related to epithelial differentiation. Therefore, there is an urgent need of reliable human physiological cell models to study viral life cycle and cancer initiation. Conditional cell reprogramming (CCR) is a rapid and robust cell culture system, where the cells can be established from minimally invasive or noninvasive specimens and their lineage functions preserved during the long-term culture. These CR cells retain their ability to differentiate at air-liquid interface (ALI). Here, we recapitulated the applications of CR and ALI approaches in modeling host-virus interactions and viral-mediated tumorigenesis.

YAP and TAZ couple osteoblast precursor mobilization to angiogenesis and mechanoregulated bone development

bioRxiv : the preprint server for biology

2023 Jan 21

Collins, JM;Lang, A;Parisi, C;Moharrer, Y;Nijsure, MP;Kim, JHT;Szeto, GL;Qin, L;Gottardi, RL;Dyment, NA;Nowlan, NC;Boerckel, JD;
PMID: 36711590 | DOI: 10.1101/2023.01.20.524918

Endochondral ossification requires coordinated mobilization of osteoblast precursors with blood vessels. During adult bone homeostasis, vessel adjacent osteoblast precursors respond to and are maintained by mechanical stimuli; however, the mechanisms by which these cells mobilize and respond to mechanical cues during embryonic development are unknown. Previously, we found that deletion of the mechanoresponsive transcriptional regulators, YAP and TAZ, from Osterix-expressing osteoblast precursors and their progeny caused perinatal lethality. Here, we show that embryonic YAP/TAZ signaling couples vessel-associated osteoblast precursor mobilization to angiogenesis in developing long bones. Osterix-conditional YAP/TAZ deletion impaired endochondral ossification in the primary ossification center but not intramembranous osteogenesis in the bone collar. Single-cell RNA sequencing revealed YAP/TAZ regulation of the angiogenic chemokine, Cxcl12, which was expressed uniquely in vessel-associated osteoblast precursors. YAP/TAZ signaling spatially coupled osteoblast precursors to blood vessels and regulated vascular morphogenesis and vessel barrier function. Further, YAP/TAZ signaling regulated vascular loop morphogenesis at the chondro-osseous junction to control hypertrophic growth plate remodeling. In human cells, mesenchymal stromal cell co-culture promoted 3D vascular network formation, which was impaired by stromal cell YAP/TAZ depletion, but rescued by recombinant CXCL12 treatment. Lastly, YAP and TAZ mediated mechanotransduction for load-induced osteogenesis in embryonic bone.

Detection of Epstein-Barr virus encoded RNA in fixed cells and tissues using CRISPR/Cas-mediated RCasFISH

Analytical biochemistry

2021 Apr 26

Chen, K;Wang, M;Zhang, R;Li, J;
PMID: 33915117 | DOI: 10.1016/j.ab.2021.114211

Identification of Epstein-Barr virus (EBV)-infected cells is critical for the diagnosis and clinical management of EBV-associated diseases. EBV-encoded RNA (EBER) located in the nucleus is a reliable marker due to its high levels of expression and inherent stability in tissue specimens. EBER in situ hybridization has long been the gold standard for detecting tumor-associated latent EBV infection and is valuable in determining the primary site and radiation fields of EBV-related malignancies. However, reliable detection is somewhat restricted by diffused signal and time-consuming procedure of this method, especially when proteins and RNA needed to be labeled simultaneously. Here, we optimized and validated our CRISPR-dCas9 mediated in situ RNA imaging tool-RCasFISH that enabled us to detect EBER rapidly and was compatible with IHC methods in fixed cells and tissue sections. Our approach could provide an attractive alternative for the molecular diagnosis of latent EBV infection.

Prognostic implications and interaction of L1 methylation and p53 expression statuses in advanced gastric cancer.

Clin Epigenetics.

2019 May 14

Shin YJ, Kim Y, Wen X, Cho NY, Lee S, Kim WH, Kang GH.
PMID: 31088544 | DOI: 10.1186/s13148-019-0661-x

Abstract

BACKGROUND:

TP53 is frequently mutated across various tissue types of cancers. In normal cells, long interspersed nuclear element-1 (LINE-1, L1) is mostly repressed by DNA methylation in its 5' untranslated region but is activated by DNA demethylation process during tumorigenesis. p53 is indispensable for maintaining genomic stability and plays its role in controlling genomic stability by repressing retrotransposon activity. However, it is unclear whether p53 regulates expression or methylation of L1 differently depending on the mutational status of TP53. Four hundred ninety cases of advanced gastric cancer (AGC) were analyzed for their statuses in p53 expression and L1 methylation using immunohistochemistry and pyrosequencing, respectively. Whether L1 methylation and expression statuses were differently affected by types of TP53 mutants was analyzed in gastric cancer cell line.

RESULTS:

By p53 immunohistochemistry, tumors were classified into 4 groups according to the intensity and extent of stained tumor nuclei. L1 methylation level was significantly higher in p53 expression group 1 than in the other groups in which L1 methylation level was similar (P < 0.001). Although L1 methylation and p53 expression statuses were associated with patient survival, multivariate analysis revealed that L1 methylation was an independent prognostic parameter. In in vitro analysis of AGS cells with the introduction of wild type or mutant types of TP53, L1 methylation level and activity were different depending on types of TP53 mutation.

CONCLUSIONS:

Findings suggest that L1 methylation level is affected by TP53 mutation status; although, L1 methylation status was an independent prognostic parameter in patients with AGC. Further study is required to elucidate the mechanism of how wild type or mutant p53 affects L1 activity and methylation status of L1 CpG island.

Brain matters: unveiling the distinct contributions of region, age, and sex to glia diversity and CNS function

Acta neuropathologica communications

2023 May 22

Seeker, LA;Bestard-Cuche, N;Jäkel, S;Kazakou, NL;Bøstrand, SMK;Wagstaff, LJ;Cholewa-Waclaw, J;Kilpatrick, AM;Van Bruggen, D;Kabbe, M;Baldivia Pohl, F;Moslehi, Z;Henderson, NC;Vallejos, CA;La Manno, G;Castelo-Branco, G;Williams, A;
PMID: 37217978 | DOI: 10.1186/s40478-023-01568-z

The myelinated white matter tracts of the central nervous system (CNS) are essential for fast transmission of electrical impulses and are often differentially affected in human neurodegenerative diseases across CNS region, age and sex. We hypothesize that this selective vulnerability is underpinned by physiological variation in white matter glia. Using single nucleus RNA sequencing of human post-mortem white matter samples from the brain, cerebellum and spinal cord and subsequent tissue-based validation we found substantial glial heterogeneity with tissue region: we identified region-specific oligodendrocyte precursor cells (OPCs) that retain developmental origin markers into adulthood, distinguishing them from mouse OPCs. Region-specific OPCs give rise to similar oligodendrocyte populations, however spinal cord oligodendrocytes exhibit markers such as SKAP2 which are associated with increased myelin production and we found a spinal cord selective population particularly equipped for producing long and thick myelin sheaths based on the expression of genes/proteins such as HCN2. Spinal cord microglia exhibit a more activated phenotype compared to brain microglia, suggesting that the spinal cord is a more pro-inflammatory environment, a difference that intensifies with age. Astrocyte gene expression correlates strongly with CNS region, however, astrocytes do not show a more activated state with region or age. Across all glia, sex differences are subtle but the consistent increased expression of protein-folding genes in male donors hints at pathways that may contribute to sex differences in disease susceptibility. These findings are essential to consider for understanding selective CNS pathologies and developing tailored therapeutic strategies.

Telocytes: Active Players in the Rainbow Trout (Oncorhynchus mykiss) Intestinal Stem-Cell Niche

Animals : an open access journal from MDPI

2021 Dec 30

Verdile, N;Pasquariello, R;Cardinaletti, G;Tibaldi, E;Brevini, TAL;Gandolfi, F;
PMID: 35011180 | DOI: 10.3390/ani12010074

In order to improve the sustainability of trout farming, it is essential to develop alternatives to fish-based meals that prevent intestinal disorders and support growth performances. Therefore, an accurate knowledge of intestinal morphology and physiology is desirable. We previously described the epithelial component of the intestinal stem-cell (ISC) niche in rainbow trout (Oncorhynchus mykiss), which is one of the most successfully farmed species and a representative model of the salmonids family. This work aims to expand that knowledge by investigating the niche stromal components that contribute to intestinal homeostasis. We analyzed samples belonging to five individuals collected from a local commercial farm. Histological and ultrastructural studies revealed peculiar mesenchymal cells adjacent to the epithelium that generated an intricate mesh spanning from the folds' base to their apex. Their voluminous nuclei, limited cytoplasm and long cytoplasmic projections characterized them as telocytes (TCs). TEM analysis showed the secretion of extracellular vesicles, suggesting their functional implication in cell-to-cell communication. Furthermore, we evaluated the localization of well-defined mouse TC markers (pdgfrα and foxl1) and their relationship with the epithelial component of the niche. TCs establish a direct connection with ISCs and provide short-range signaling, which also indicates their key role as the mesenchymal component of the stem-cell niche in this species. Interestingly, the TC distribution and gene-expression pattern in rainbow trout closely overlapped with those observed in mice, indicating that they have the same functions in both species. These results substantially improve our understanding of the mechanisms regulating intestinal homeostasis and will enable a more detailed evaluation of innovative feed effects.

	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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