RESUMO
Group 3 innate lymphoid cells (ILC3s) are mediators of intestinal immunity and barrier function. Recent studies have investigated the role of the mammalian target of rapamycin complex (mTOR) in ILC3s, whereas the mTORC1-related mechanisms and crosstalk between mTORC1 and mTORC2 involved in regulating ILC3 homeostasis remain unknown. In this study, we found that mTORC1 but not mTORC2 was critical in ILC3 development, IL-22 production, and ILC3-mediated intestinal homeostasis. Single-cell RNA sequencing revealed that mTORC1 deficiency led to disruption of ILC3 heterogeneity, showing an increase in differentiation into ILC1-like phenotypes. Mechanistically, mTORC1 deficiency decreased the expression of NFIL3, which is a critical transcription factor responsible for ILC3 development. The activities of both mTORC1 and mTORC2 were increased in wild-type ILC3s after activation by IL-23, whereas inhibition of mTORC1 by Raptor deletion or rapamycin treatment resulted in increased mTORC2 activity. Previous studies have demonstrated that S6K, the main downstream target of mTORC1, can directly phosphorylate Rictor to dampen mTORC2 activity. Our data found that inhibition of mTORC1 activity by rapamycin reduced Rictor phosphorylation in ILC3s. Reversing the increased mTORC2 activity via heterozygous or homozygous knockout of Rictor in Raptor-deleted ILC3s resulted in severe ILC3 loss and complete susceptibility to intestinal infection in mice with mTORC1 deficiency (100% mortality). Thus, mTORC1 acts as a rheostat of ILC3 heterogeneity, and mTORC2 protects ILC3s from severe loss of cells and immune activity against intestinal infection when mTORC1 activity is diminished.
Assuntos
Imunidade Inata , Linfócitos , Camundongos , Animais , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Proteína Companheira de mTOR Insensível à Rapamicina/metabolismo , Proteína Regulatória Associada a mTOR/genética , Fatores de Transcrição/metabolismo , Sirolimo/farmacologia , Mamíferos/metabolismoRESUMO
BACKGROUND: The tumor suppressor gene CDH1 is critical for intercellular adhesion. In our previous work, we reported a nonfunctional CDH1 transcript that lacks the final 83 base pairs of exon 8 (1054del83). In this work, we probed the role of histone epigenetic modifications as well as DNA methylation in selection of this isoform. METHODS: RT-qPCR was used to detect CDH1 RNA expression. Methylation of CDH1 was analyzed by bisulphite sequencing PCR. ChIP assay was performed to show histones level. Cell lines were treated with DNA methyltransferase inhibitor AZA, HDAC inhibitor TSA, or siRNA oligonucleotides to test regulation of CDH1 splicing. RESULTS: Greater CDH1 1054del83 transcripts were observed in gastric cancer (GC) cell lines than human gastric mucosal epithelial cell line GES-1. All the cell lines showed significant methylation pattern at the CpG sites of CDH1 exon 8. AZA treatment did not influence selection of 1054del83 transcripts. A significant decrease in acetylation for histones H3 and H4K16Ac in an internal region of the CDH1 gene surrounding the alternative exon 8 were detected in GC cell lines. Treatment with TSA preferentially expressed the correctly spliced transcript and not the exon 8 skipped aberrant transcripts, showing that histone acetylation was involved in the splicing regulation. SiRNA-mediated knockdown of SETD2 (The specific methyltransferase of H3K36) decreased exclusion of exon 8, suggesting that the presence of this mark correlates with increased skipping of the final 83 base pairs of CDH1 exon 8. However, CDH1 splicing was not affected by SRSF2 knockdown. CONCLUSIONS: H3K36me3 correlates with increased skipping of the final 83 base pairs of CDH1 exon 8. Histone acetylation was involved in the splicing regulation as well.
Assuntos
Processamento Alternativo/genética , Caderinas/genética , Metilação de DNA/genética , Epigênese Genética/genética , Neoplasias Gástricas/genética , Antígenos CD , Sequência de Bases , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Éxons/genética , Histonas/genética , Humanos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
OBJECTIVE: To explore the diagnosis and treatment of testicular intratubular seminoma. METHODS: One case of testicular intratubular seminoma was diagnosed with testicular biopsy. Epididymal sperm was aspirated and intracytoplasmic sperm injection ( ICSI) was performed. And local radiotherapy was conducted on the bilateral testes after fertilization. RESULTS: The result of testicular biopsy revealed bilateral testicular intratubular seminoma. Large numbers of sperms were found in the epididymal aspirate. ICSI did not succeed for the first time. The second ICSI succeeded. The local radiotherapy by 60Co had been conducted on the bilateral testes, and testicular tumor didn't develop further. CONCLUSION: Testicular intratubular seminoma is a type of intratubular germ cell neoplasia, with no clinical manifestations, and usually found in testicular biopsy. Earlier management promises better prognosis.