Probes for EPHB2

Cancer stem cells (CSCs) are believed to be responsible for tumor growth, invasion, and metastasis. Submucosal invasion, which greatly enhances metastasis risk, is a critical step in gastric cancer (GC) progression. To identify stem cell-related markers associated with submucosal invasion and lymph node (LN) metastasis in GCs, we investigated the expression of candidate CSC markers (CD133, CD44, and ALDH1A) and intestinal stem cell (ISC) markers (EPHB2, OLFM4, and LGR5) in early GCs that manifested submucosal invasion. We discovered that EPHB2 and LGR5 expression was frequently confined to the basal area of the lamina propria (basal pattern) in mucosal cancer, and the proportion of stem cell marker-positive cells substantially increased during submucosal invasion. CD44 expression showed a focal pattern, ALDH1A was predominantly expressed diffusely, and there was no expansion of CD44 or ALDH1A expression in the submucosal cancer cells. Unexpectedly, no CSC markers showed any associations with LN metastasis, and only loss of EPHB2 expression was associated with increased LN metastasis. Treatment of RSPO2, a niche factor, along with Wnt 3a, to GC cells led to increased EPHB2 and LGR5 mRNA levels. RNA in situ hybridization confirmed specific RSPO2 expression in the smooth muscle cells of the muscularis mucosa, suggesting that RSPO2 is responsible for the increased expression of ISC markers in GC cells at the basal areas. In summary, no stem cell markers were associated with increased LN metastasis in early GCs. Conversely, isolated EPHB2 expression was associated with lower LN metastasis. EPHB2 and LGR5 showed a basal distribution pattern along with enhanced expression in submucosal invading cells in early GCs, which was induced by a niche factor, RSPO2, from the muscularis mucosa.

Abstract Aims Intestinal stem cell (ISC) markers such as LGR5, ASCL2, EPHB2 and OLFM4 and their clinical implications have been extensively studied in colorectal cancers (CRCs). However, little is known about their expression in precancerous lesions of CRCs. Here, we investigated the expression and distribution of ISC markers in serrated polyps and conventional adenomas. Methods and results RT-PCR analysis revealed that all ISC markers were significantly upregulated in conventional adenomas with low grade dysplasia (CALGs) compared with other lesions. RNA in situ hybridization confirmed that CALGs exhibited strong and diffuse expression of all ISC markers, which indicate a stem cell-like phenotype. However, normal colonic mucosa hyperplastic polyps and sessile serrated adenomas harbored LGR5+ cells that were confined to the crypt base and demonstrated an organized expression of ISC markers. Notably, in traditional serrated adenomas, expression of LGR5 and ASCL2 was localized to the ectopic crypts as in the normal crypts, but expression of EPHB2 and OLFM4 was distributed in a diffuse manner, which is suggestive of a progenitor-like features. Conclusions The expression and distribution profile of ISC markers possibly provides insights into the organization of stem and progenitor-like cells in each type of precancerous lesion of CRC

Social behavior starts with dynamic approach prior to the final consummation. The flexible processes ensure mutual feedback across social brains to transmit signals. However, how the brain responds to the initial social stimuli precisely to elicit timed behaviors remains elusive. Here, by using real-time calcium recording, we identify the abnormalities of EphB2 mutant with autism-associated Q858X mutation in processing long-range approach and accurate activity of prefrontal cortex (dmPFC). The EphB2-dependent dmPFC activation precedes the behavioral onset and is actively associated with subsequent social action with the partner. Furthermore, we find that partner dmPFC activity is responsive coordinately to the approaching WT mouse rather than Q858X mutant mouse, and the social defects caused by the mutation are rescued by synchro-optogenetic activation in dmPFC of paired social partners. These results thus reveal that EphB2 sustains neuronal activation in the dmPFC that is essential for the proactive modulation of social approach to initial social interaction.

Gastric intestinal metaplasia (IM) is a highly prevalent preneoplastic lesion; however, the molecular mechanisms regulating its development remain unclear. We have previously shown that a population of cells expressing the intestinal stem cell (ISC) marker LGR5 increases remarkably in IM. In this study, we further investigated the molecular characteristics of these LGR5+ cells in IM by examining the expression profile of several ISC markers. Notably, we found that ISC markers-including OLFM4 and EPHB2-are positively associated with the CDX2 expression in non-tumorous gastric tissues. This finding was confirmed in stomach lesions with or without metaplasia, which demonstrated that OLFM4 and EPHB2 expression gradually increased with metaplastic progression. Moreover, RNA in situ hybridization revealed that LGR5+ cells coexpress several ISC markers and remained confined to the base of metaplastic glands, reminiscent to that of normal intestinal crypts, whereas those in normal antral glands expressed none of these markers. Furthermore, a large number of ISC marker-expressing cells were diffusely distributed in gastric adenomas, suggesting that these markers may facilitate gastric tumorigenesis. In addition, Barrett's esophagus (BE)-which is histologically similar to intestinal metaplasia-exhibited a similar distribution of ISC markers, indicating the presence of a stem cell population with intestinal differentiation potential. In conclusion, we identified that LGR5+ cells in gastric IM and BE coexpress ISC markers, and exhibit the same expression profile as those found in normal intestinal crypts. Taken together, these results implicate an intestinal-like stem cell population in the pathogenesis of IM, and provide an important basis for understanding the development and maintenance of this disease.

Current therapeutic strategies for treating nonalcoholic steatohepatitis (NASH) have failed to alleviate liver fibrosis, which is a devastating feature leading to hepatic dysfunction. Here, we integrated single-nucleus transcriptomics and epigenomics to characterize all major liver cell types during NASH development in mice and humans. The bifurcation of hepatocyte trajectory with NASH progression was conserved between mice and humans. At the nonalcoholic fatty liver (NAFL) stage, hepatocytes exhibited metabolic adaptation, whereas at the NASH stage, a subset of hepatocytes was enriched for the signatures of cell adhesion and migration, which were mainly demarcated by receptor tyrosine kinase ephrin type B receptor 2 (EphB2). EphB2, acting as a downstream effector of Notch signaling in hepatocytes, was sufficient to induce cell-autonomous inflammation. Knockdown of Ephb2 in hepatocytes ameliorated inflammation and fibrosis in a mouse model of NASH. Thus, EphB2-expressing hepatocytes contribute to NASH progression and may serve as a potential therapeutic target.