Development of a 45K pepper GBTS liquid-phase gene chip and its application in genome-wide association studies.

Introduction: Pepper (Capsicum spp.) is a vegetable that is cultivated globally and has undergone extensive domestication, leading to a significant diversification in its agronomic traits. With the advancement of genomics in pepper and the reduction in sequencing costs, the high-throughput detection of single nucleotide polymorphisms (SNPs) and small insertions-deletions (indels) has become increasingly critical for analyzing pepper germplasms and improving breeding programs. As a result, there is a pressing need for a cost-effective, high-throughput, and versatile technique suitable for both foreground and background selection in pepper breeding. Methods: In the present study, Python-based web scraping scripts were utilized to systematically extract data from published literatures and relevant sequence databases focusing on pepper genomes. Subsequent to data extraction, SNPs and indels were meticulously identified and filtered. This process culminated in the delineation of core polymorphic sites, which were instrumental in the development of specific probes. Following this, comprehensive phenotypic and genotypic analyses were conducted on a diverse collection of 420 pepper germplasms. Concurrently, a genome-wide association study (GWAS) was conducted to elucidate the genetic determinants of helical fruit shape in peppers. Results: In this study, a 45K pepper Genotyping-By-Target-Sequencing (GBTS) liquid-phase gene chip was developed on the GenoBaits platform. This chip is composed of 45,389 probes, of which 42,535 are derived from core polymorphic sites (CPS) in the background genetic landscape, while 2,854 are associated with foreground agronomic traits, spanning across 43 traits. The CPS probes are spaced at an average interval of 68 Kb. We have assessed the performance of this chip on 420 pepper germplasms, with successful capture of target DNA fragments by 45,387 probes. Furthermore, the probe capture ratio surpassed 70% in 410 of the 420 germplasms tested. Using this chip, we have efficiently genotyped 273 germplasms for spiciness levels and elucidated the genetic relationships among 410 pepper germplasms. Our results allowed for precise clustering of sister lines and C. chinense germplasms. In addition, through a GWAS for helical fruit shape, we identified three quantitative trait loci (QTLs): heli2.1, heli11.1, and heli11.2. Within the heli11.1 QTL, a gene encoding the tubulin alpha chain was identified, suggesting its potential role in the helical growth pattern of pepper fruits. Discussion: In summary, the 45K pepper GBTS liquid-phase gene chip offers robust detection of polymorphic sites and is a promising tool for advancing research into pepper germplasm and the breeding of new pepper varieties.

The Therapeutic Target of IBD and the Mechanism of Dipyridamole in Treating IBD Explored by Geo Gene Chips, Network Pharmacology, and Molecular Docking.

BACKGROUND AND AIMS: Inflammatory Bowel Disease (IBD) is a refractory disease with repeated attacks, and there is no accurate treatment target at present. Dipyridamole, a phosphodiesterase (PDE) inhibitor, has been proven to be an effective treatment for IBD in a pilot study. This study explored the therapeutic target of IBD and the pharmacological mechanism of dipyridamole for the treatment of IBD. MATERIALS AND METHODS: The candidate targets of dipyridamole were obtained by searching the pharmMapper online server and Swiss Target Prediction Database. The IBD-related targets were selected from four GEO chips and three databases, including Genecards, DisGeNET, and TTD database. A protein-protein interaction (PPI) network was constructed, and the core targets were identified according to the topological structure. KEGG and GO enrichment analysis and BioGPS location were performed. Finally, molecular docking was used to verify dipyridamole and the hub targets. RESULTS: We obtained 112 up-regulated genes and 157 down-regulated genes, as well as 105 composite targets of Dipyridamole-IBD. Through the PPI network analysis, we obtained the 7 hub targets, including SRC, EGFR, MAPK1, MAPK14, MAPK8, PTPN11, and LCK. The BioGPS showed that these genes were highly expressed in the immune system, digestive system, and endocrine system. In addition, the 7 hub targets had good intermolecular interactions with dipyridamole. The therapeutic effect of dipyridamole on IBD may involve immune system activation and regulation of inflammatory reactions involved in the regulation of extracellular matrix, perinuclear region of cytoplasm, protein kinase binding, and positive regulation of programmed cell death through cancer pathway (proteoglycans in cancer), lipid metabolism, Ras signaling pathway, MAPK signaling pathway, PI3K-AKT signaling pathway, Th17 cell differentiation, and other cellular and innate immune signaling pathways. CONCLUSION: This study predicted the therapeutic target of IBD and the molecular mechanism of dipyridamole in treating IBD, providing a new direction for the treatment of IBD and a theoretical basis for further research.

Differentially expressed long non-coding RNAs and mRNAs of cadmium exposure on learning disability of offspring rats.

BACKGROUND: Cadmium (Cd) exposure has been found to have detrimental effects on the development of the central nervous system and cognitive ability in children. However, there is ongoing debate regarding the impact of maternal Cd exposure on the cognitive ability of offspring. In this study, we aimed to investigate the mechanisms underlying the influence of maternal Cd exposure on the cognitive ability of offspring rats. METHODS: Here, we constructed a model of cadmium poisoning in first-generation rats through gavage. The cognitive and memory abilities of its offspring were evaluated by water maze experiment. Then, we used the gene chip to find out the key genes, and we performed qRT-PCR detection of these genes. Subsequently, enrichment analysis was employed to identify pathways. Finally, we constructed a co-expression network consisting of LncRNAs and mRNAs to elucidate the biological functions and regulatory mechanisms of LncRNAs. RESULTS: The results of the water maze trial demonstrated that the offspring of rats exposed to cadmium in the first generation had reduced cognitive and memory abilities. Through an analysis of gene expression in the hippocampus of the cadmium-treated rats' offspring and the control group, we identified a correlation between the islet secretion pathway and the cognitive impairment observed in the offspring. Utilizing various algorithms, we identified Cpa1 and Prss1 as potential key genes associated with the cognitive impairment caused by cadmium. The results of qRT-PCR demonstrated a decrease in the expression levels of these genes in the hippocampus of the cadmium-treated rats' offspring. In addition, in the co-expression network, we observed that Cpa1 was co-expressed with 11 LncRNAs, while Prss1 was associated with 4 unexplored LncRNAs. Furthermore, we conducted an analysis to examine the relationship between Cpa1, Prss1-related transcription factors, and LncRNAs. CONCLUSION: Overall, this study provides novel insights into the molecular effects of first generation Cd exposure on the cognitive ability of offspring. The target genes and signaling pathways investigated in this study could serve as potential targets for improving neurodevelopment and cognitive ability in children.

Advances in the application of molecular diagnostic techniques for the detection of infectious disease pathogens (Review).

Infectious diseases are a major global cause of morbidity and mortality, seriously affecting public health and socioeconomic stability. Since infectious diseases can be caused by a wide variety of pathogens with similar clinical manifestations and symptoms that are difficult to accurately distinguish, selecting the appropriate diagnostic techniques for the rapid identification of pathogens is crucial for clinical disease diagnosis and public health management. However, traditional diagnostic techniques have low detection rates, long detection times and limited automation, which means that they do not meet the requirements for rapid diagnosis. Recent years have seen continuous developments in molecular detection technology, which has a higher sensitivity and specificity, shorter detection time and increased automation, and performs an important role in the early and rapid detection of infectious disease pathogens. The present study summarizes recent progress in molecular diagnostic technologies such as PCR, isothermal amplification, gene chips and high­throughput sequencing for the detection of infectious disease pathogens, and compares the technical principles, advantages and disadvantages, applicability and costs of these diagnostic techniques.

Analysis of circular RNA expression profile of pathological bone formation in ankylosing spondylitis.

OBJECTIVE: To screen and analyze the function of specific CircRNAs involved in pathological bone formation in patients with ankylosing spondylitis (AS). METHODS: From September 2019 to October 2020, hip capsule tissues obtained from 3 patients with AS developed hip joint fusion and 3 patients with femoral neck fracture (FNF) were obtained. The circular RNA expressions of hip capsule were analyzed by Arraystar CircRNA chip. qRT-PCR analysis wan performed to identify the expression patterns of differentially expression CircRNAs. RESULTS: Our findings showed that there were 25 up-regulated and 39 down-regulated differential CircRNAs. Among these CircRNAs, we screened 10 highest up-regulatedCircRNAs and 13 lowest down-regulated CircRNAs (Fold Change≥2,P<0.05). In further verification analysis, hsa_circ_0067103, hsa_circ_0004496, and hsa_circ_0002649, ACTG1 were significantly upregulated, while hsa_circ_0020273, hsa_circ_0005699, and hsa_circ_0048764 were markly downregulated in AS tissue than FNF controls. CONCLUSION: The expression of CircRNAs involved of pathological bone formation in AS were significantly different from those of control group. These differentially expressed Circular RNAs may be closely related to the occurrence and development of pathological bone formation in AS.

Application and prospect of gene chip in genetic breeding of livestock and poultry.

Gene chip is a high-throughput technique for detecting specific DNA sequences by DNA or DNA-RNA complementary hybridization, among which SNP genotyping chips have been widely employed in the animal genetics and breeding, and have made great achievements in cattle (Bos taurus), pigs (Sus scrofa), sheep (Caprinae), chickens (Gallus gallus) and other livestock. However, genomic selection applied in production merely uses genomic information and cannot fully explain the molecular mechanism of complex traits genetics, which limits the accuracy of genomic selection. With the continuous progresses in epigenetic research, the development of commercial methylation chips and the application of the epigenome-wide association study (EWAS), DNA methylation has been extensively used to draw the causal connections between genetics and phenotypes. In the future, it is hopefully expected to develop methylation chips customized for livestock and poultry and explore methylation sites significantly related to economic traits of livestock and poultry through EWAS thereby extending the understanding of causal variation of complex traits. Combining methylation chips and SNP chips, we can capture the epigenomic and genomic information of livestock and poultry, interpret genetic variation more precisely, improve the accuracy of genome selection, and promote the fine evolution of molecular genetic breeding of livestock and poultry. In this review, we summarize the application of SNP chips and depict the prospects of the application of methylation chips in livestock and poultry. This review will provide valuable insights for further application of gene chips in farm animal breeding.

Molecular Methods for Identification and Quantification of Foodborne Pathogens.

Foodborne pathogens that enter the human food chain are a significant threat worldwide to human health. Timely and cost-effective detection of them became challenging for many countries that want to improve their detection and control of foodborne illness. We summarize simple, rapid, specific, and highly effective molecular technology that is used to detect and identify foodborne pathogens, including polymerase chain reaction, isothermal amplification, loop-mediated isothermal amplification, nucleic acid sequence-based amplification, as well as gene chip and gene probe technology. The principles of their operation, the research supporting their application, and the advantages and disadvantages of each technology are summarized.

Gene expression profiling in nucleus pulposus of human ruptured lumbar disc herniation.

Purpose: To examine the differences in gene expression between ruptured and non-ruptured nucleus pulposus tissues of the intervertebral discs using gene chip technology. Methods: A total of 8 patients with nucleus pulposus from a lumbar disc herniation (LDH) undergoing discectomy in our hospital were selected, including 4 ruptured and 4 non-ruptured herniated nucleus pulposus cases. Total RNA was extracted from cells by using TRIzol reagent. Nucleus pulposus cDNA probes of the two groups were obtained by the single marker method and hybridized with a human gene expression profiling chip (Agilent). The fluorescence signal images were scanned by a laser, and the obtained genes were analyzed by bioinformatics. Results: There were 75 differentially expressed genes with more than 2-fold-changes, of which 56 were up-regulated and 19 were down-regulated. The differential expression of THSD7A, which was up-regulated 18 times, was the most significant, followed by CCL5, AQP3 and SDC4. Conclusion: THSD7A can be used as a characteristic differentially expressed gene in human ruptured nucleus pulposus. Moreover, CCL5, AQP3 and SDC4 may improve the chemotaxis of stem cell migration for self-healing of damaged disc tissue, increase water uptake by nucleus accumbens cells, and inhibit the inflammatory response, thus delaying the process of intervertebral disc degeneration.

Bioinformatics analysis of potential key genes and pathways in neonatal necrotizing enterocolitis.

OBJECTIVE: To detect differentially expressed genes in patients with neonatal necrotizing enterocolitis (NEC) by bioinformatics methods and to provide new ideas and research directions for the prevention, early diagnosis and treatment of NEC. METHODS: Gene chip data were downloaded from the Gene Expression Omnibus database. The genes that were differentially expressed in NEC compared with normal intestinal tissues were screened with GEO2R. The functions, pathway enrichment and protein interactions of these genes were analyzed with DAVID and STRING. Then, the core network genes and significant protein interaction modules were detected using Cytoscape software. RESULTS: Overall, a total of 236 differentially expressed genes were detected, including 225 upregulated genes and 11 downregulated genes, and GO and KEGG enrichment analyses were performed. The results indicated that the upregulated differentially expressed genes were related to the dimerization activity of proteins, while the downregulated differentially expressed genes were related to the activity of cholesterol transporters. KEGG enrichment analysis revealed that the differentially expressed genes were significantly concentrated in metabolism, fat digestion and absorption pathways. Through STRING analysis, 9 key genes in the protein network interaction map were identified: EPCAM, CDH1, CFTR, IL-6, APOB, APOC3, APOA4, SLC2A and NR1H4. CONCLUSION: Metabolic pathways and biological processes may play important roles in the development of NEC. The screening of possible core targets by bioinformatics is helpful in clarifying the pathogenesis of NEC at the gene level and in providing references for further research.

Development of a quadruple PCR-based gene microarray for detection of vaccine and wild-type classical swine fever virus, African swine fever virus and atypical porcine pestivirus.

BACKGROUND: Classical swine fever (CSF), African swine fever (ASF), and atypical porcine pestivirus (APPV) are acute, virulent, and contagious viral diseases currently hampering the pig industry in China, which result in mummification or stillbirths in piglets and mortality in pigs. Diagnostic assays for the differentiation of infection and vaccination of CSFV, in addition to the detection of ASFV and APPV, are urgently required for better prevention, control, and elimination of these viral diseases in China. METHODS: A quadruple PCR-based gene microarray assay was developed in this study to simultaneously detect wild-type and vaccine CSFV strains, ASFV and APPV according to their conserved regions. Forty-two laboratory-confirmed samples, including positive samples of 10 other swine viral diseases, were tested using this assay to confirm its high specificity. RESULTS: This assay's limit of detections (LODs) for the wild-type and vaccine CSFV were 6.98 and 6.92 copies/µL. LODs for ASFV and APPV were 2.56 × 10 and 1.80 × 10 copies/µL, respectively. When compared with standard RT-PCR or qPCR for CSFV (GB/T 26875-2018), ASFV (MARR issue No.172), or APPV (CN108611442A) using 219 clinical samples, the coincidence was 100%. The results showed that this assay with high sensitivity could specifically distinguish ASFV, APPV, and CSFV, including CSFV infection and immunization. CONCLUSION: This assay provides a practical, simple, economic, and reliable test for the rapid detection and accurate diagnosis of the three viruses and may have good prospects for application in an epidemiological investigation, prevention, and control and elimination of these three diseases.

Identification of Novel Imatinib-Resistant Genes in Gastrointestinal Stromal Tumors.

Gastrointestinal stromal tumors (GISTs) are common ICC precursor sarcomas, which are considered to be a potential malignant mesenchymal tumor driven by specific KIT or PDGFRA signals in the gastrointestinal tract. The standard treatment for GIST without metastasis is surgical resection. GIST with metastasis is usually treated with tyrosine kinase inhibitors (TKIs) only but cannot be cured. The TKI imatinib is the main drug of GIST drug therapy. In adjuvant therapy, the duration of imatinib adjuvant therapy is 3 years. It has been proved that imatinib can improve the overall survival time (OS). However, many GIST patients develop drug resistance due to the long-term use of imatinib. We were forced to look for new strategies to treat GIST. The purpose of the current academic work is to study the drug-resistant genes of imatinib and their potential mechanisms. A total of 897 differentially expressed genes (DEGs) were found between imatinib-sensitive cell line GIST882 and imatinib-resistant cell line GIST430 by RNA sequencing (RNA-seq). After analyzing the DEGs, 10 top genes were selected (NDN, FABP4, COL4A1, COLEC11, MEG3, EPHA3, EDN3, LMO3, RGS4, and CRISP2). These genes were analyzed by RT-PCR, and it was confirmed that the expression trend of FABP4, COL4A1, and RGS4 in different imatinib-resistant cell lines was in accord with the GEO database. It is suggested that these genes may play a potential role in the clinical diagnosis and treatment of imatinib resistance in GIST.

Octacosanol Modifies Obesity, Expression Profile and Inflammation Response of Hepatic Tissues in High-Fat Diet Mice.

The incidence of obesity has increased significantly on account of the alterations of living habits, especially changes in eating habits. In this study, we investigated the effect of octacosanol on lipid lowering and its molecular mechanism. High-fat diet (HFD)-induced obesity mouse model was used in the study. Thirty C57BL/6J mice were divided into control, HFD, and HFD+Oct groups randomly, and every group included ten mice. The mice of HFD+Oct group were intragastrically administrated 100 mg/kg/day of octacosanol. After 10 weeks for treatment, our results indicated that octacosanol supplementation decreased the body, liver, and adipose tissues weight of HFD mice; levels of TC, TG, and LDL-c were reduced in the plasma of HFD mice; and level of HDL-c were increased. H&E staining indicated that octacosanol supplementation reduces the size of fat droplets of hepatic tissues and adipose cells comparing with the HFD group. Gene chip analysis found that octacosanol regulated 72 genes involved in lipid metabolism in the tissues of liver comparing to the HFD group. IPA pathway network analysis indicated that PPAR and AMPK may play a pivotal role in the lipid-lowering function of octacosanol. Real-time quantitative PCR and Western blot showed that the octacosanol supplementation caused change of expression levels of AMPK, PPARs, FASN, ACC, SREBP-1c, and SIRT1, which were closely related to lipid metabolism. Taken together, our results suggest that octacosanol supplementation exerts a lipid-decreasing effect in the HFD-fed mice through modulating the lipid metabolism-related signal pathway.

Advances in multiplex molecular detection technologies for harmful algae.

As the eutrophication of natural water bodies becomes more and more serious, the frequency of outbreaks of harmful algal blooms (HABs) mainly formed by harmful algae also increases. HABs have become a global ecological problem that poses a serious threat to human health and food safety. Therefore, it is extremely important to establish methods that can rapidly detect harmful algal species for early warning of HABs. The traditional morphology-based identification method is inefficient and inaccurate. In recent years, the rapid development of molecular biology techniques has provided new ideas for the detection of harmful algae and has become a research hotspot. The current molecular detection methods for harmful algal species mainly include fluorescence in situ hybridization, sandwich hybridization, and quantitative PCR (qPCR), but all of these methods can only detect single harmful algal species at a time. The establishment of methods for the simultaneous detection of multiple harmful algal species has become a new trend in the development of molecular detection technology because various harmful algal species may coexist in the natural water environment. The established molecular techniques for multiple detections of harmful algae mainly include gene chip, multiplex PCR, multiplex qPCR, massively parallel sequencing, antibody chip, and multiple isothermal amplification. This review mainly focuses on the principles, advantages and disadvantages, application progress, and application prospects of these multiple detection technologies, aiming at providing effective references not only for the fisheries but also for economic activities, environment, and human health.

[Bioinformatics Analysis of Microarray Data in Myelodysplastic Syndrome Based on Gene Expression Omnibus Database].

OBJECTIVE: To identify the key genes and explore mechanisms in the development of myelodysplastic syndrome (MDS) by bioinformatics analysis. METHODS: Two cohorts profile datasets of MDS were downloaded from Gene Expression Omnibus (GEO) database. Differentially expressed gene (DEG) was screened by GEO2R, functional annotation of DEG was gained from GO database, gene ontology (GO) enrichment analysis was performed via Kyoto Encyclopedia of Genes and Genomes (KEGG) database, and key genes were screened by Matthews correlation coefficient (MCC) based on STRING database. RESULTS: There were 112 DEGs identified, including 85 up-regulated genes and 27 down-regulated genes. GO enrichment analysis showed that biological processes were mainly enriched in immune response, etc, cellular component in cell membrane, etc, and molecular function in protein binding, etc. KEGG signaling pathway analysis showed that main gene enrichment pathways were primary immunodeficiency, hematopoietic cell lineage, B cell receptor signaling pathway, Hippo signaling pathway, and asthma. Three significant modules were screened by Cytoscape software MCODE plug-in, while 10 key node genes (CD19, CD79A, CD79B, EBF1, VPREB1, IRF4, BLNK, RAG1, POU2AF1, IRF8) in protein-protein interaction (PPI) network were screened based on STRING database. CONCLUSION: These screened key genes and signaling pathways are helpful to better understand molecular mechanism of MDS, and provide theoretical basis for clinical targeted therapy.

A Comparative Study of Systems Pharmacology and Gene Chip Technology for Predicting Targets of a Traditional Chinese Medicine Formula in Primary Liver Cancer Treatment.

Background: The systems pharmacology approach is a target prediction model for traditional Chinese medicine and has been used increasingly in recent years. However, the accuracy of this model to other prediction models is yet to be established. Objective : To compare the systems pharmacology modelwithexperimental gene chip technology by using these models to predict targets of a traditional Chinese medicine formulain the treatment of primary liver cancer. Methods: Systems pharmacology and gene chip target predictions were performed for the traditional Chinese medicine formula ZhenzhuXiaojiTang (ZZXJT). A third square alignment was performed with molecular docking. Results: Identification of systems pharmacology accounted for 17% of targets, whilegene chip-predicted outcomes accounted for 19%.Molecular docking showed that the top ten targets (excludingcommon targets) of the system pharmacology model had better binding free energies than the gene chip model using twocommon targets as a benchmark. For both models, the core drugs predictions were more consistent than the core small molecules predictions. Conclusion:In this study, the identified targets of systems pharmacology weredissimilar to those identified by gene chip technology; whereas the core drug and small molecule predictions were similar.

Expression of IER3 in hepatocellular carcinoma: clinicopathology, prognosis, and potential regulatory pathways.

Background: Immediate early response 3 (IER3) is correlated to the prognosis of several cancers, but the precise mechanisms underlying the regulation by IER3 of the occurrence and development of hepatocellular carcinoma (HCC) remain unknown. Methods: The expression level of IER3 was examined by using in-house immunohistochemistry (IHC), public gene chip, and public RNA-sequencing (RNA-seq). The standardized mean difference (SMD) was calculated to compare the expression levels of IER3 between HCC patients and controls. The summary receiver operating characteristics (sROC) was plotted to comprehensively understand the discriminatory capability of IER3 between HCC and non-HCC group. The Kaplan-Meier curves and the combined hazard ratios (HRs) were used to determine the prognostic value of IER3 in HCC. Moreover, differentially expressed genes (DEGs) and co-expression genes (CEGs) were used to explored the molecular mechanisms of IER3 underlying HCC. hTFtarget was used to predict the transcription factors (TFs) of IER3. The binding site of TFs and the IER3 promoter region was forecasted using the JASPAR website. The relevant ChIP-seq data were used to determine whether TF peaks were present in the IER3 transcription initiation. Results: A significantly increased expression of IER3 protein was found in HCC tissue relative to non-HCC tissue as detected by IHC (p < 0.001). Compared to 1,263 cases of non-HCC tissues, IER3 in 1483 cases of HCC tissues was upregulated (SMD = 0.42, 95% confidence interval [CI] [0.09-0.76]). The sROC showed that IER3 had a certain ability at differentiating HCC tissues (area under the curve (AUC) = 0.65, 95% CI [0.61-0.69]). Comprehensive analysis of the effect of IER3 on the prognosis of patients with HCC demonstrated that higher IER3 expression was associated with poor prognosis in HCC (HRs = 1.30, 95% CI [1.03-1.64]). Pathway enrichment analysis revealed that IER3-related genes were mostly enriched in the PI3K-Akt signaling pathway, cancer-related signaling pathways, the p53 signaling pathway, and other signaling pathways. Regulatory factor X5 (RFX5) was identified as a possible regulator of IER3-related TF. Conclusion: IER3 may be a potential prognostic marker for HCC. The molecular mechanisms of IER3 in HCC warrant further study.

Exploring the pathogenesis of type 2 diabetes mellitus intestinal damp-heat syndrome and the therapeutic effect of Gegen Qinlian Decoction from the perspective of exosomal miRNA.

ETHNOPHARMACOLOGICAL RELEVANCE: Diabetes is a common, complex, chronic metabolic disease. A randomized, double-blind, placebo-parallel controlled clinical study has shown that Gegen Qinlian Decoction (GQD) can reduce glycosylated hemoglobin in type 2 diabetes mellitus (T2DM) intestinal damp-heat syndrome patients in a dose-dependent manner. AIM: To explore the pathogenesis of T2DM intestinal damp-heat syndrome and the therapeutic effect of GQD from the perspective of exosomal microRNA (miRNA). METHODS: Eligible patients were selected and treated with GQD for 3 months to evaluate their clinical efficacy. Effective cases were matched with healthy volunteers, and saliva samples were collected. Exosomal miRNA was extracted from saliva and analyzed by chip sequencing. Subsequently, the function of the differential gene and the signal transduction pathway were analyzed using bioinformatics technology. Finally, three target miRNAs were randomly selected from the T2DM group/healthy group, and two target miRNAs in the T2DM before treatment/after treatment group were randomly selected for qPCR verification. Finally, we conducted a correlation analysis of the miRNAs and clinical indicators. The registration number for this research is ChiCTR-IOR-15006626. RESULTS: (1) The expression of exosomal miRNA chips showed that there were 14 differentially expressed miRNAs in the T2DM group/healthy group, and 26 differentially expressed miRNAs in the T2DM before treatment/after treatment group. (2) Enrichment results showed that in the T2DM group/healthy group, it was primarily related to cell development, body metabolism, TGF-ß, and ErbB signaling pathways. In the T2DM before treatment/after treatment group, it was mainly related to cellular metabolic regulation processes, and insulin, Wnt, and AMPK signaling pathways. (3) The qPCR verification showed that the expressions of hsa-miR-9-5p, hsa-miR-150-5p, and hsa-miR-216b-5p in the T2DM group was higher (P<0.05). Following GQD treatment, hsa-miR-342-3p and hsa-miR-221-3p were significantly downregulated (P<0.05). (4) hsa-miR-9-5p was positively correlated with BMI (P<0.05), and hsa-miR-150-5p was positively correlated with total cholesterol and triglycerides (P<0.05). The GQD efficacy-related gene hsa-miR-342-3p was positively correlated with the patient's initial blood glucose level (P<0.05), and hsa-miR-221-3p was positively correlated with total cholesterol and triglycerides (P<0.05). CONCLUSION: The exosomal miRNA expression profile and signaling pathways related to T2DM intestinal damp-heat syndrome and the efficacy of GQD were established, which provides an alternative strategy for precision traditional Chinese medicine treatment.

[Impact of Cadmium on the expressions of piRNAs in the rat testis].

OBJECTIVE: To study the impact of cadmium (Cd) on the expressions of PIWI-interacting RNAs (piRNA) in the rat testis and its possible action mechanism. METHODS: Twelve 6-week-old SD rats were randomly divided into a Cd-exposure and a control group, the former gavaged with CdCl2 at 3 mg/kg/d and the latter with normal saline, all for 28 successive days. Then the testicular tissues were collected from the rats, sperm concentration and motility were obtained by computer-assisted sperm analysis (CASA), and piRNA sequencing was performed using the gene chip, followed by bioinformatics analysis of differentially expressed piRNAs. RESULTS: Compared with the controls, the rats in the Cd-exposure group showed significantly decreased sperm concentration and motility (P < 0.05). The expressions of 272 piRNAs were up-regulated and 402 down-regulated after 28 days of Cd exposure, and 4 of the up-regulated piRNAs were consistent with the results of gene chip verification. Bioinformatics analysis showed that the 4 up-regulated piRNA target genes were involved in 50 biological processes, such as negative regulation of apoptosis, positive regulation of gene expression and positive regulation of GTPase activity, and mainly concentrated in 13 signaling pathways including transcription dysregulation, calcium and mitogen-activated protein kinase signaling pathways in cancer. Among them, PIRNA-DQ765261 had a binding site with Bcl-2. CONCLUSION: Cadmium can induce changes in the expressions piRNAs in the rat testicular tissue, and some piRNAs may be involved in the autophagy and apoptosis of sperm. Bcl-2 may be the target of PIRNA-DQ765261.

Clinical application of gene chip technology in diagnosis and treatment of silicosis complicated with mycobacterial infection / 环境与职业医学

Background Gene chip technology has been increasingly used in the diagnosis and treatment of common tuberculosis. However, its role in the diagnosis and treatment of silicosis complicated with mycobacterial infection remains unclear. Objective To evaluate the application value of gene chip technology in the diagnosis and treatment of silicosis complicated with mycobacterial infection. Methods From January 2019 to June 2021, 197 silicosis patients suspected to be complicated with mycobacterial infection in Quanzhou First Hospital Affiliated to Fujian Medical University were enrolled in this study. The etiology evaluation for the 197 patients was conducted by acid-fast staining of sputum smear (sputum smear method), culture of Mycobacterium tuberculosis of sputum (sputum culture method), and gene chip technology of bronchoalveolar lavage fluid (BALF); and for 80 patients among them, acid-fast staining of BALF (BALF smear method) and culture of Mycobacterium tuberculosis of BALF (BALF culture method) were additionally performed. The positive rates and consistency were assessed using intraclass correlation coefficient (ICC). Test for Mycobacterium tuberculosis drug resistance mutation gene was added for patients with Mycobacterium tuberculosis complex by BALF gene chip technology. Results The average age of the 197 patients was (53.1±9.1) years, and the average dust exposure time was (21.1±9.4) years, including 192 males and 5 females. There were 8 cases with stage I silicosis, 17 cases with stage II silicosis, and 172 cases with stage III silicosis. Among them, 11.2% were positive for sputum smear; 24.4% were positive for sputum culture, and 36.0% were positive by gene chip of BALF. The difference between the three methods was statistically significant (P<0.05). The result of consistency test for the three methods showed that the ICC was 0.539 (P<0.001). Among the 80 patients, there was a significant difference in the positive rates of the five methods (χ2=25.23, P<0.001). The results of Bonferroni test showed statistically significant pair-wise differences between BALF culture method and sputum smear method, BALF culture method and BALF smear method, BALF gene chip method and sputum smear method, BALF gene chip method and BALF smear method (P<0.05), while there were no statistically significant differences between the other pairs (P>0.05). The result of consistency test for the five methods showed that the ICC was 0.586 (P<0.001). Among the 71 BALF gene chip positive cases, 59 cases reported positive Mycobacterium tuberculosis complex (17 cases were positive in the first-line anti-tuberculosis resistance test, and 2 cases were found positive quinolone resistance gene in the second-line anti-tuberculosis resistance test), and received regular anti-tuberculosis treatment, among them 45 cases improved and 14 cases were stable; 12 cases reported non-tuberculous mycobacteria cases, among them 5 cases received anti-non-tuberculous mycobacteria treatment (4 cases improved and 1 case was stable), and 7 cases with mild symptoms did not receive anti-non-tuberculous mycobacteria treatment. Conclusion Compared with sputum smear, sputum culture, and other traditional methods, gene chip technology of BALF can improve the positive rate of pathogenic diagnosis of silicosis complicated with mycobacterial infection, and can also quickly identify whether it is non-tuberculous mycobacteria infection or drug-resistant Mycobacterium tuberculosis infection, which is helpful to adjust treatment as soon as possible.

Bioinformatics Analysis of Microarray Data in Myelodysplastic Syndrome Based on Gene Expression Omnibus Database / 中国实验血液学杂志

OBJECTIVE@#To identify the key genes and explore mechanisms in the development of myelodysplastic syndrome (MDS) by bioinformatics analysis.@*METHODS@#Two cohorts profile datasets of MDS were downloaded from Gene Expression Omnibus (GEO) database. Differentially expressed gene (DEG) was screened by GEO2R, functional annotation of DEG was gained from GO database, gene ontology (GO) enrichment analysis was performed via Kyoto Encyclopedia of Genes and Genomes (KEGG) database, and key genes were screened by Matthews correlation coefficient (MCC) based on STRING database.@*RESULTS@#There were 112 DEGs identified, including 85 up-regulated genes and 27 down-regulated genes. GO enrichment analysis showed that biological processes were mainly enriched in immune response, etc, cellular component in cell membrane, etc, and molecular function in protein binding, etc. KEGG signaling pathway analysis showed that main gene enrichment pathways were primary immunodeficiency, hematopoietic cell lineage, B cell receptor signaling pathway, Hippo signaling pathway, and asthma. Three significant modules were screened by Cytoscape software MCODE plug-in, while 10 key node genes (CD19, CD79A, CD79B, EBF1, VPREB1, IRF4, BLNK, RAG1, POU2AF1, IRF8) in protein-protein interaction (PPI) network were screened based on STRING database.@*CONCLUSION@#These screened key genes and signaling pathways are helpful to better understand molecular mechanism of MDS, and provide theoretical basis for clinical targeted therapy.