Endogenous hydrogen sulfide promotes human preimplantation embryonic development by regulating metabolism-related gene expression.

Hydrogen sulfide (H2S) as an endogenous gaseous signaling molecule had been proved to play a vital role in gametes physiology, covering meiosis, maturation and aging. However, little is known about H2S involvement in embryonic development. The present study explored the positive effect of H2S on human early embryonic development. Results validated that the two H2S producing enzymes, CBS and CSE mRNA and proteins were identified in donated human cleavage and blastocyst-stage embryos. The l-cysteine incubation produced endogenous H2S in human blastocysts. NaHS positively affected in vitro blastulation. Single-cell RNA-seq analysis identified 228 differentially expressed genes (DEGs) after NaHS treatment versus the control. The Gene Ontology (GO) enrichment analysis of DEGs showed that genes for protein modification and metabolism were significantly enriched in the NaHS treatment group. For the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, 2-oxocarboxylic acid metabolism, glycosaminoglycan biosynthesis-keratan sulfate, steroid biosynthesis, carbon metabolism, and biosynthesis of amino acids were significantly enriched. Six DEGs, including Neural EGFL like 1 (NELL1), aconitase 1 (ACO1), phosphoglycerate mutase 1 (PGAM1), TP53 induced glycolysis regulatory phosphatase (TIGAR), UDP-GlcNAc:betaGal beta-1,3-N-acetylglucosaminyltransferase 2 (B3GNT2), and carbohydrate Sulfotransferase 4 (CHST4) were validate by real-time RT-PCR. These findings suggest that H2S is a positive regulator of early embryonic development and may alter the transcription of embryonic genes for protein modification and metabolism in human embryos.

Identification of a distinct luminal subgroup diagnosing and stratifying early stage prostate cancer by tissue-based single-cell RNA sequencing.

BACKGROUND: The highly intra-tumoral heterogeneity and complex cell origination of prostate cancer greatly limits the utility of traditional bulk RNA sequencing in finding better biomarker for disease diagnosis and stratification. Tissue specimens based single-cell RNA sequencing holds great promise for identification of novel biomarkers. However, this technique has yet been used in the study of prostate cancer heterogeneity. METHODS: Cell types and the corresponding marker genes were identified by single-cell RNA sequencing. Malignant states of different clusters were evaluated by copy number variation analysis and differentially expressed genes of pseudo-bulks sequencing. Diagnosis and stratification of prostate cancer was estimated by receiver operating characteristic curves of marker genes. Expression characteristics of marker genes were verified by immunostaining. RESULTS: Fifteen cell groups including three luminal clusters with different expression profiles were identified in prostate cancer tissues. The luminal cluster with the highest copy number variation level and marker genes enriched in prostate cancer-related metabolic processes was considered the malignant cluster. This cluster contained a distinct subgroup with high expression level of prostate cancer biomarkers and a strong distinguishing ability of normal and cancerous prostates across different pathology grading. In addition, we identified another marker gene, Hepsin (HPN), with a 0.930 area under the curve score distinguishing normal tissue from prostate cancer lesion. This finding was further validated by immunostaining of HPN in prostate cancer tissue array. CONCLUSION: Our findings provide a valuable resource for interpreting tumor heterogeneity in prostate cancer, and a novel candidate marker for prostate cancer management.

Mutational Pattern in Multiple Pulmonary Nodules Are Associated With Early Stage Lung Adenocarcinoma.

The clinical significance of mutation in multiple pulmonary nodules is largely limited by single gene mutation-directed analysis and lack of validation of gene expression profiles. New analytic strategy is urgently needed for comprehensive understanding of genomic data in multiple pulmonary nodules. In this study, we performed whole exome sequencing in 16 multiple lung nodules and 5 adjacent normal tissues from 4 patients with multiple pulmonary nodules and decoded the mutation information from a perspective of cellular functions and signaling pathways. Mutated genes as well as mutation patterns shared in more than two lesions were identified and characterized. We found that the number of mutations or mutated genes and the extent of protein structural changes caused by different mutations is positively correlated with the degree of malignancy. Moreover, the mutated genes in the nodules are associated with the molecular functions or signaling pathways related to cell proliferation and survival. We showed a developing pattern of quantity (the number of mutations/mutated genes) and quality (the extent of protein structural changes) in multiple pulmonary nodules. The mutation and mutated genes in multiple pulmonary nodules are associated with cell proliferation and survival related signaling pathways. This study provides a new perspective for comprehension of genomic mutational data and might shed new light on deciphering molecular evolution of early stage lung adenocarcinoma.

iTRAQ-based proteomic analysis of human umbilical vein endothelial cells with platelet endothelial aggregation receptor-1 knockdown.

The disorders of hemostasis and coagulation were believed to be the main contributors to the pathogenesis of pulmonary thromboembolism (PTE), and platelets are the basic factors regulating hemostasis and coagulation and play important roles in the process of thrombosis. This study investigated the proteome of human umbilical vein endothelial cells (HUVECs) with platelet endothelial aggregation receptor-1 (PEAR1) knockdown using the isobaric tags for relative and absolute quantitation (iTRAQ) method and analyzed the role of differential abundance proteins (DAPs) in the regulation of platelets aggregation. Our results showed that the conditioned media-culturing HUVECs with PEAR1 knockdown partially suppressed the adenosine diphosphate (ADP)-induced platelet aggregation. The proteomics analysis was performed by using the iTRAQ technique, and a total of 215 DAPs (124 protein was upregulated and 91 protein were downregulated) were identified. The Gene Ontology (GO) enrichment analysis showed that proteins related to platelet α granule, adenosine triphosphate metabolic process, and endocytosis were significantly enriched. Further, the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis also identified the significant enrichment of endocytosis-related pathways. The real-time polymerase chain reaction assay confirmed that the expression of P2Y12 , mitochondrial carrier 2, NADH dehydrogenase (ubiquinone) iron-sulfur protein 3, and ubiquinol-cytochrome c reductase hinge protein are significantly downregulated in the HUVECs with PEAR1 knockdown. In conclusion, our in vitro results implicated that DAPs induced by PEAR1 knockdown might contribute to the platelet aggregation. Proteomic studies by employing GO enrichment and KEGG pathway analysis suggested that the potential effects of DAPs on platelet aggregation may be linked to the balance of ADP synthesis or degradation in mitochondria.

Hydrogen Sulfide Facilitates Vaginal Lubrication by Activation of Epithelial ATP-Sensitive K(+) Channels and Cystic Fibrosis Transmembrane Conductance Regulator.

INTRODUCTION: Hydrogen sulfide (H2S) plays a large role in female and male sexual responses characterized by a smooth muscle relaxant effect. Moreover, H2S is a novel pro-secretory neuromodulator that modulates epithelial ion transport. However, whether H2S has a role in regulating vaginal epithelial ion transport and fluid secretion has not been extensively studied. AIM: To identify the effects of H2S on vaginal epithelial ion transport and lubrication in an exploratory investigation. METHODS: The mRNA, protein expression, and localization of cystathionine γ-lyase (CSE) and H2S production in vaginal epithelium were examined by reverse transcriptase polymerase chain reaction, Western blot, H2S synthesizing activity assay, and immunohistochemistry, respectively. The effect of H2S on vaginal epithelial ion transport, vaginal fluid secretion, and ionic concentration was investigated using a short-circuit current (ISC), a measurement of vaginal lubrication, and ion chromatography, respectively. MAIN OUTCOME MEASURES: The mRNA, protein expression, and localization of CSE, H2S formation, changes of ISC responses, vaginal lubrication, and K(+) and Cl(-) concentrations were studied. RESULTS: CSE mRNA and protein were predominantly expressed in vaginal epithelium. Sodium hydrosulfide hydrate (NaHS) caused concentration-dependent changes in ISC across isolated rat vaginal epithelium, which consisted of an initial decrease phase and then an increase phase. The increase phase in ISC was mainly Cl(-) dependent and abolished by cystic fibrosis transmembrane conductance regulator inhibitor, whereas the decrease phase was sensitive to the adenosine triphosphate-sensitive K(+) (KATP) channel blocker. Furthermore, intravaginal treatment of NaHS significantly enhanced vaginal lubrication in vivo, and this effect was prevented by cystic fibrosis transmembrane conductance regulator and KATP channel inhibitors. In addition, the ionic concentrations of K(+) and Cl(-) in rat vaginal fluid were significantly increased by NaHS treatment. CONCLUSION: The CSE-H2S pathway participates in the regulation of vaginal epithelial K(+) and Cl(-) ion transport to modulate lumen fluid secretion.

Aberrant activation of hedgehog pathway in nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is a very common head and neck cancer in southern china. Despite advances in surgical and chemotherapeutic approaches, its prognosis is still not promising. Hedgehog signaling pathway was reported to be involved in a number of cancers including head and neck. However, it remains unclear regarding the role of this pathway in NPC. By real-time PCR, we found Ptch1, Smo, and Gli-1 were expressed in all human nasopharyngeal epithelial tissues and cell lines. Compared with nasopharyngeal normal epithelial tissues, the mRNA expression level of Gli-1 was higher in carcinoma and nasopharyngitis (NPI) epithelial tissues. While compared with nasopharyngitis epithelia, the mRNA expression level of Ptch1 was lower in carcinoma epithelia and normal epithelia. The expressions of Smo mRNA were not significantly different among these epithelial tissues. Immunohistochemistry analysis revealed that the expression level of Gli-1 was higher in NPC than NPI. Thus, our data indicated that aberrant activation of hedgehog pathway in NPC. Furthermore, blocking the pathway with cyclopamine inhibited the proliferation of NPC epithelia cell lines. In addition, blockade of the pathway in three NPC cell lines with cyclopamine-induced tumor cell apoptosis. The transcription of hedgehog target genes also is inhibited by cyclopamine. These data suggested that hedgehog pathway may sustain nasopharyngeal tumor growth. Our data demonstrated that hedgehog signaling pathway was involved in NPC pathogenesis and might be a novel therapeutic target for NPC.