Objective To map the genome-wide distribution profile of histone H3K27me3 modification in diffuse gastric cancer tissues,identify target genes regulated by H3K27me3,and primarily explore the potential mechanism of its modification reprogramming in the occurrence and development of the tumor.Methods Normal gastric mucosal tissues and diffuse gastric cancer tissues were harvested from the patients who underwent examinations or treatments in the departments of gastroenterology and gastrointestinal surgery of our medical center between 2021 and 2023.There were 14 patients in the normal group(6 males and 8 females,average age of 46 years)and 14 patients in the gastric cancer group(8 males and 6 females,average age of 63 years).Cleavage under target and tagmentation(CUT&Tag)technology was employed to capture genomic regions modified by H3K27me3,and analyze the reprogramming characteristics of these modifications.RNA sequencing data,data from high-throughput chromosome conformation capture(Hi-C)technology,and publicly available single-cell data were integrated to investigate the target genes regulated by the reprogramming of H3K27me3 modifications in diffuse gastric cancer cells.Results The quality of the CUT&Tag and RNA sequencing data met the standards required for subsequent analysis.Histone H3K27me3 modifications in normal gastric mucosa and diffuse gastric cancer tissues were primarily distributed in distal intergenic regions and intronic regions.In gastric cancer tissues,compared to normal tissues,there was significant reprogramming of H3K27me3 modifications,characterized by a marked reduction in overall H3K27me3 signal intensity.The loss of 2 912 H3K27me3 signal peaks might lead to the up-regulation of 822 tumor-associated genes.Among them,56 genes displayed the most significant up-regulation(fold change in signal intensity≥2,P<0.05),with notable enrichment in the mammalian target of rapamycin complex 1(mTORC1)signaling pathway.Specifically,the methionine transporter SLC7A5 and the cystine transporter SLC7A11 were found to have the highest expression levels in gastric cancer tissues.Single-cell data revealed that the abnormal overexpression of SLC7A11 in diffuse gastric cancer was primarily observed in tumor epithelial cells.Further validation using public data and immunohistochemical experiments confirmed the elevated expression of SLC7A11 in diffuse gastric cancer,which is associated with poor prognosis in gastric cancer patients.Conclusion The reprogramming of histone H3K27me3 modification is an important epigenetic characteristic in diffuse gastric cancer.Loss of H3K27me3 signal peaks may up-regulate the expression of SLC7A11 in diffuse gastric cancer cells,and thereby promote tumor progression.

Objective To draw the genome-wide distribution and remodeling characteristics of H3K27me3 silencers in signet-ring cell carcinoma of the stomach(SRCC)through epigenetic sequencing technology,and to investigate their roles in transcriptional regulation in order to elucidate the regulatory mechanism of SRCC malignant progression.Methods The study was conducted on 35 gastric samples obtained by gastroendoscopic biopsy(15 normal and 20 SRCC tissues)from Department of Gastroenterology of Army Medical Center of PLA between January 2021 and December 2023.Multi-omics analyses,including assay for transposase-accessible chromatin with high-throughput sequencing(ATAC-seq),cleavage under targets and tagmentation(CUT&Tag)and transcriptome sequencing(RNA-seq),were performed to identify chromatin accessibility,H3K27me3 silencer regions,and transcriptional changes,with aid of Illumina NovaSeq 6000.H3K27me3 related differentially expressed genes(|Log2FC|>1,FDR<0.05)were screened using DESeq2.Gene Ontology(GO)analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis were employed to analyze the enrichment function,and Homer was employed to identify transcription factor motifs.A regulatory network was constructed using Cytoscape,and then validated using immunohistochemistry to explore its regulatory mechanism.Results H3K27me3 silencers were primarily located in distal intergenic regions(37.06%)in SRCC.Compared with the normal tissues,SRCC showed a significant reduction in H3K27me3 silencer signals(95%CI:1.34～2.30,P=0.007)with 6 257 lost sites(FDR<0.01).Integrating CUT&Tag and RNA-seq revealed 380 up-regulated immune-related genes,particularly in T cell receptor signaling(OR=4.2,95%CI:2.8～6.3,P=0.002).Immunohistochemistry confirmed elevated expression of transcription factor EHF(P<0.05).Conclusion There is the remodeling of H3K27me3 silencers in SRCC,and EHF may potentially play a crucial role in the SRCC malignant progression.

Background:The core circadian clock gene Bmal1 has been shown to be involved in the formation of visceral sensitization in irritable bowel syndrome(IBS)by affecting the tryptophan hydroxylase 1(TPH1)-5-hydroxytryptamine(5-HT)pathway,but the exact mechanism of its regulation is unknown.Aims:To investigate the molecular mechanism by which Bmal1 regulates the TPH1-5-HT pathway through the clock-controlled gene Piezo1.Methods:Dexamethasone was used to synchronize the expression of the circadian clock genes in RIN-14B cells.Bmal1 expression was up-regulated or down-regulated in RIN-14B cells by plasmid and siRNA transfection of the enterochromaffin cell(EC)model.The expression levels of target genes and proteins were detected by immunofluorescence staining,RT-qPCR,and Western blotting.5-HT content was detected by ELISA method.Results:(1)This study was the first to report the oscillation characteristics of RIN-14B circadian clock genes in EC model,among which the oscillation of Bmal1 was the most significant.Immunofluorescence showed that RIN-14B cells expressed CGA,Bmal1 and Piezo1.(2)After transfected with the Bmal1 overexpression plasmid,the mRNA and protein expression of Bmal1 were significantly up-regulated in RIN-14B cells compared with the negative control group(all P<0.001);while transfected with Bmal1 siRNA significantly decreased the mRNA and protein expression of Bmal1 in RIN-14B cells compared with the negative control group(all P<0.05).(3)After transfected with Bmal1 overexpression plasmid,the mRNA and protein expression of Piezo1,the protein expression of TPH1,and the intracellular content of 5-HT were significantly increased(all P<0.051).(4)After transfected with Bmal siRNA,mRNA expression of Piezo1 and TPH1 in RIN-14B cells was significantly down-regulated(all P<0.05),and the protein expression of Piezo1,TPH1,the intracellular 5-HT content tended to be decreased by 21%,31%,and 10%,respectively.Conclusions:RIN-14B cells have the characteristics of rhythmic oscillation of circadian clock genes,and Bmal1 overexpression and underexpression EC models can be successfully established by using RIN-14B cells.Overexpression and underexpression of Bmal1 can lead to significant changes in the clock-controlled Piezo1-TPH1-5-HT signaling pathway,suggesting that Bmal1 can be expressed by clock-control signals through the Piezo1-TPH1 pathway.This suggests that Bmal1 may be involved in the development of visceral hypersensitivity in IBS through regulation of 5-HT synthesis by the clock-controlled gene Piezo1.

Background:The core circadian clock gene Bmal1 has been shown to be involved in the formation of visceral sensitization in irritable bowel syndrome(IBS)by affecting the tryptophan hydroxylase 1(TPH1)-5-hydroxytryptamine(5-HT)pathway,but the exact mechanism of its regulation is unknown.Aims:To investigate the molecular mechanism by which Bmal1 regulates the TPH1-5-HT pathway through the clock-controlled gene Piezo1.Methods:Dexamethasone was used to synchronize the expression of the circadian clock genes in RIN-14B cells.Bmal1 expression was up-regulated or down-regulated in RIN-14B cells by plasmid and siRNA transfection of the enterochromaffin cell(EC)model.The expression levels of target genes and proteins were detected by immunofluorescence staining,RT-qPCR,and Western blotting.5-HT content was detected by ELISA method.Results:(1)This study was the first to report the oscillation characteristics of RIN-14B circadian clock genes in EC model,among which the oscillation of Bmal1 was the most significant.Immunofluorescence showed that RIN-14B cells expressed CGA,Bmal1 and Piezo1.(2)After transfected with the Bmal1 overexpression plasmid,the mRNA and protein expression of Bmal1 were significantly up-regulated in RIN-14B cells compared with the negative control group(all P<0.001);while transfected with Bmal1 siRNA significantly decreased the mRNA and protein expression of Bmal1 in RIN-14B cells compared with the negative control group(all P<0.05).(3)After transfected with Bmal1 overexpression plasmid,the mRNA and protein expression of Piezo1,the protein expression of TPH1,and the intracellular content of 5-HT were significantly increased(all P<0.051).(4)After transfected with Bmal siRNA,mRNA expression of Piezo1 and TPH1 in RIN-14B cells was significantly down-regulated(all P<0.05),and the protein expression of Piezo1,TPH1,the intracellular 5-HT content tended to be decreased by 21%,31%,and 10%,respectively.Conclusions:RIN-14B cells have the characteristics of rhythmic oscillation of circadian clock genes,and Bmal1 overexpression and underexpression EC models can be successfully established by using RIN-14B cells.Overexpression and underexpression of Bmal1 can lead to significant changes in the clock-controlled Piezo1-TPH1-5-HT signaling pathway,suggesting that Bmal1 can be expressed by clock-control signals through the Piezo1-TPH1 pathway.This suggests that Bmal1 may be involved in the development of visceral hypersensitivity in IBS through regulation of 5-HT synthesis by the clock-controlled gene Piezo1.

We developed a high-efficiency microfluidic chip for extracting exosomes from human plasma. We collected peripheral blood from normal human, designed and fabricated a microfluidic chip based on nanoporous membrane and agarose gel electrophoresis to isolate exosomes. The extracted exosomes were characterized by transmission electron microscopy, nanosight and Western blotting, the morphology, concentration and particle size of exosomes were identified and analyzed. Meanwhile, we used ultracentrifugation and microfluidic chip to isolate exosomes separately. The particle size and concentration of the exosomes extracted by two methods were compared and analyzed, and their respective extraction efficiency was discussed. Finally, the expression level of miRNA-21 in exosomes was analyzed by RT-PCR. The microfluidic chip isolated (in 1 hour) high-purity exosomes with size ranging from 30-200 nm directly from human plasma, allowing downstream exosomal miRNA analysis. By comparing with ultracentrifugation, the isolation yield of microfluidic chip was 3.80 times higher than ultracentrifugation when the volume of plasma sample less than 100 μL. The optimized parameters for exosome isolation by gel electrophoresis microfluidic chip were: voltage: 100 V; concentration of agarose gel: 1.0%; flow rate of injection pump: 0.1 mL/h. The gel electrophoresis microfluidic chips could rapidly and efficiently isolate the exosomes, showing great potential in the research of exosomes and cancer biomarkers.
Exosomes ; Humans ; MicroRNAs/genetics* ; Microfluidics ; Plasma ; Ultracentrifugation

To investigate roles of nitric oxide (NO) signal in accumulations of phenolic acids in abscisic.acid (ABA)-induced Salvia miltiorrhiza hairy roots, S. miltiorrhiza hairy roots were treated with different concentrations of sodium nitroprusside (SNP)-an exogenous NO donor, for 6 days, and contents of phenolic acids in the hairy roots are determined. Then with treatment of ABA and NO scavenger (2-(4-carboxy-2-phenyl)-4,4,5,5-tetramethylimidazoline-1- oxyl-3-oxide, c-PTIO) or NO synthase inhibitor (NG-nitro-L-arginine methyl ester, L-NAME), contents of phenolic acids and expression levels of three key genes involved in phenolic acids biosynthesis were detected. Phenolic acids production in S. miltiorrhiza hairy roots was most significantly improved by 100 µmoL/L SNP. Contents of RA and salvianolic acid B increased by 3 and 4 folds. ABA significantly improved transcript levels of PAL (phenylalanine ammonia lyase), TAT (tyrosine aminotransferase) and RAS (rosmarinic acid synthase), and increased phenolic acids accumulations. However, with treatments of ABA+c-PTIO or ABA+L-NAME, accumulations of phenolic acids and expression levels of the three key genes were significantly inhibited. Both NO and ABA can increase accumulations of phenolic acids in S. miltiorrhiza hairy roots. NO signal probably mediates the ABA-induced phenolic acids production.
Abscisic Acid ; pharmacology ; Benzofurans ; metabolism ; Free Radical Scavengers ; pharmacology ; Hydroxybenzoates ; metabolism ; Nitric Oxide ; metabolism ; Phenylalanine Ammonia-Lyase ; metabolism ; Plant Roots ; metabolism ; Salvia miltiorrhiza ; metabolism ; Tyrosine Transaminase ; metabolism

BACKGROUND:Surgical treatment is commonly used for decompressing the spinal cord in multilevel cervical spondylotic myelopathy, but the optimum anterior cervical reconstructive method has not been determined. OBJECTIVE:To compare and analyze the biomechanical characteristics of four anterior cervical reconstructive techniques in the surgical management of multilevel cervical spondylotic myelopathy utilizing the three-dimensional finite element model. METHODS:A three-dimensional finite element model of an intact C2–7 segment was developed and validated based on healthy adult male CT images. Four additional models were developed from the fusion model, including anterior cervical discectomy and fusion, anterior cervical corpectomy and fusion, anterior cervical hybrid decompression and fusion and anterior cervical discectomy and fusion with Cage alone. Von Mises stresses on the plate and the disc of adjacent levels (C2/3, C6/7) were comparatively analyzed. RESULTS AND CONCLUSION:(1) The C2/3 disc stress in flexion, extension, lateral bending and rotation condition was significantly higher than the C6/7 disc in 4 anterior cervical reconstructive techniques, and the adjacent disc stress (C2/3, C6/7) was lowest in the anterior cervical discectomy and fusion with Cage alone, and highest in the anterior cervical corpectomy and fusion. (2) The stress at the plate-screw interface was highest in the anterior cervical corpectomy and fusion, and lowest in the anterior cervical discectomy and fusion. (3) In conclusion, among the four anterior cervical reconstructive techniques for multilevel cervical spondylotic myelopathy, the anterior cervical discectomy and fusion with Cage alone makes little effect on the adjacent disc stress, which might reduce the incidence of adjacent segment disease after fusion. However, the accompanying risk of the increased incidence of cage subsidence should never be neglected.

OBJECTIVETo explore the effects of early intervention of acupuncture on urinary function reconstruction of neurogenic bladder after spinal cord injury, so as to find the optimal treatment timing of acupuncture.

METHODSFifty-three patients of neurogenic bladder after spinal cord injury were randomly divided into a prior-to-catheter group (25 cases) and a posterior-to-catheter group (28 cases). For the prior-to-catheter group, one week before the removal of catheter, acupoints on the abdomen and back such as Guanyuan (CV 4), Zhongji (CV 3), Baliao (Shangliao (BL 31), Ciliao (BL 32), Zhongliao (BL 33), Xialiao (BL 34), Jiaji (EX-B 2) and acupoints at distal end such as Zusanli (ST 36) and Sanyinjiao (SP 6) were selected; after the catheter removal, simple method. of water column was used to measure bladder capacity pressure to classify the type of bladder, and then different acupuncture manipulations were given; intermittent catheterization was performed if condition was allowed. The posterior-to-catheter group applied identical treatment after removal of catheter as the prior-to-catheter group. The treatment was given 5 times per week; after one-month treatment, five aspects including the time of first effective urination, time of establishing reflex urination, average residual volume of urine, time of residual urine less than 100 mL and quality of life (QOL) were evaluated.

RESULTSAfter treatment, the time of first effective urination, time of establishing reflex urination, average residual volume of urine and time of residual urineless than 100 mL in the prior-to-catheter group were all superior to those in the posterior-to-catheter group (all P<0. 05) ; the score of QOL was improved in two groups after treatment (both P<0. 01), but the difference between two groups was not statistically significant (P>0. 05); each index of urination function was not significant among patients with different injured segments of spinal cord (all P>0. 05).

CONCLUSIONThe early intervention of acupuncture (prior to catheter) has obvious improving function on establishing balanced bladder in patients with neurogenic bladder after spinal cord injury, which is superior to acupuncture intervention after removal of catheter, however, the effects of different injured segments on establishing balanced bladder are not different.

Acupuncture Points ; Acupuncture Therapy ; Adolescent ; Adult ; Aged ; Female ; Humans ; Male ; Middle Aged ; Spinal Cord Injuries ; complications ; Treatment Outcome ; Urinary Bladder ; physiopathology ; Urinary Bladder, Neurogenic ; etiology ; physiopathology ; therapy ; Urination ; Young Adult

Objective: To examine the therapeutic effects of dual dorsolateral prefrontal repetitive transcranial magnetic stimulation (rTMS) on auditory hallucinations experienced by schizophrenic patients. Methods: A double-blind randomized controlled study was conducted. Totally 25 schizophrenic patients met DSM-Ⅳ criteria for schizophrenia and had anditory hallucination with stable antipsychotic drugs treatment were recruited. Subjects were grouped into active treatment subgroup and sham subgroup randomly. Measurements including the Positive and Negative Symptom Scale (PANSS) for clinical symptom and the Treatment Emergent Symptom Scale (TESS) for side-effects were evaluated Results: Totally 25 subjects completed the trial. The treatment efficiency of active treatment group was higher than that of sham group (75% vs. 31%) .No overt side effects of rTMS were observed. Conclusion: It is effective for rTMS to treat schizophrenia with auditory hallucination at 1Hz. rTMS may be a new selection of schizophrenia treatment.

OBJECTIVETo compare therapeutic effects of red-hot needle therapy and filiform needle therapy on nodules of breast of the type of stagnation of liver-qi and phlegm coagulation.

METHODSSix hundred cases were randomly divided into a treatment group and a control group, 300 cases in each group. The treatment group were treated with red-hot needle pricking at the proliferative parts and Ashi points as main, and the control group with filiform needle therapy.

RESULTSIn the treatment group, 240 cases were cured, 58 cases improved and 2 cases were not cured with an effective rate of 99.3%; and in the control group, 113 cases were cured, 165 cases improved and 22 were not cured with an effective rate of 92.7%, with a significant difference between the two groups (P = 0.001).

CONCLUSIONThe therapeutic effect of red-hot needle therapy is better than that of filiform needle therapy on nodules of breast of the type of stagnation of liver-qi and phlegm coagulation.

Acupuncture Therapy ; methods ; Adult ; Female ; Fibrocystic Breast Disease ; etiology ; therapy ; Humans ; Middle Aged