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Long noncoding RNAs (lncRNAs) are revealed to be involved in the tumorigenesis and progression of human malignancies mediated by microRNA (miRNA) via the competing endogenous RNA (ceRNA) mechanism, a newly proposed “RNA language.” However, the lncRNA-associated competing triplet (lncACT) network among ceRNA transcripts in clear cell renal cell carcinoma (ccRCC) is currently lacking. We carried out differential expression analysis to identify aberrantly expressed lncRNAs, miRNAs, and mRNAs by analyzing the RNA-seq data of 420 ccRCC tissues and 71 noncancerous kidney tissues obtained from The Cancer Genome Atlas (TCGA). Then, a ccRCC-specific ceRNA network was built using computational algorithms, including miRcode, TargetScan, miRanda, and miRTarBase. In total, 1491 dysregulated lncRNAs were found between normal renal tissues and ccRCC (fold change > 4 and false discovery rate < 0.01 ). A ceRNA network that comprised of 46 DElncRNAs, 11 DEmiRNAs, and 55 DEmRNAs was established by integrating the lncRNA/miRNA and miRNA/mRNA interactions into lncACTs. Several lncRNAs were identified to be significantly associated with clinical features of ccRCC patients. Notably, four key lncRNAs (TCL6, HOTTIP, HULC, and PCGEM1) were tightly correlated with both patients’ clinical characteristics and overall survival (log-rank P < 0.05 ), indicating their potential important roles in ccRCC. HOTTIP may be a potential prognostic and therapeutic molecular marker for ccRCC patients. Collectively, our results provide a comprehensive view of the lncRNA-associated ceRNA regulatory network for a better understanding of the mechanisms and prognosis biomarkers for ccRCC.

Multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE) are demyelinating neuroinflammatory diseases identified by the accumulation and aggregation of misfolded proteins in the brain. The Sel1L-Hrd1 complex comprising endoplasmic reticulum associated degradation (ERAD) is an ER-protein quality control system (ERQC) in the cell. Unfortunately, the contribution of ERAD to the development of these diseases has not been well explored. In this study, we used mice with a conditional deletion (KO) of Sel1L in T cells to dissect the role of ERAD on T cells and its contribution to the development of EAE. The results showed that Sel1L KO mice developed more severe EAE than the control wild type (WT) mice. Although, no obvious effects on peripheral T cells in steady state, more CD44-CD25+ double-negative stage 3 (DN3) cells were detected in the thymus. Moreover, Sel1L deficiency promoted the differentiation of Th1 and Th17 cells and upregulated the proliferation and apoptosis of CD4 T cells in vitro. Regarding the mechanism analyzed by RNA sequencing, 437 downregulated genes and 271 upregulated genes were detected in Sel1L deletion CD4 T cells, which covered the activation, proliferation, differentiation and apoptosis of these T cells. Thus, this study declared that the dysfunction of Sel1L in ERAD in T cells exacerbated the severity of EAE and indicated the important role of ERQC in maintaining immune homeostasis in the central nervous system.

Ectoderm-derived mesenchymal stem cells (EMSCs) were used as potential seed cells for bone tissue engineering to treat bone defects due to their capability of rapid proliferation and osteogenic differentiation. Sonic hedgehog (Shh) signaling was reported to play an important role in the development of bone tissue, but its role is not understood. The present study investigated the role of Shh molecule in osteogenic differentiation of rat EMSCs in vitro. Rat EMSCs were isolated form nasal respiratory mucosa and identified with immunofluorescence and analyzed with other methods, including reverse transcriptase polymerase chain reaction (qPCR) and western blotting. EMSCs expressed CD90, CD105, nestin, and vimentin. On the seventh day of osteogenic induction, expression levels of Shh and Gli1 was higher according to the result of qPCR and Western blotting. After induction for 14 days, higher alkaline phosphatase (ALP) activity and more mineralized nodules were seen in comparison to the cells that did not undergo induction. Shh signaling appears to enhance osteogenic differentiation of rat EMSCs, suggesting that Shh signaling directs the lineage differentiation of ectodermal stem cells and represents a promising strategy for skeletal tissue regeneration.

Background: ISO 15189:2012 is widely recommended for clinical laboratories. However, it has not been paid enough attention in medical laboratory education and experiments in clinical biochemistry courses. Methods/Approach: The teaching program of clinical biochemistry aims to improve students’ awareness and ability of quality management according to ISO 15189 technical requirements. It is a 4-stage experimental training program to teaching skills: "establish an overall testing process based on the patient's clinical indicator, clarifying principles, improve operational skills, review process and continuous improvement". The program was implemented in our college during the winter semesters of 2019 and 2020. 185 students participate in the program as an experimental group, other 172 students as control group adopt the conventional method. Participants completed an online survey after each class to evaluate the class. Results/Outcomes: The test group had significantly better examination scores not only in experimental operational skills (89.27±7.16 vs. 77.51±4.72, pConclusions: A 4-stage experimental training program based on ISO 15189 technical requirements combined with CBL in clinical biochemistry is an effective and acceptable teaching strategy for the conventional training program.

Epidermal growth factor receptor (EGFR) signalling and the interleukin-6 (IL-6)/signal transducer and activator of transcription 3 (STAT3) are aberrantly activated in ovarian cancer. However, inhibition of EGFR signalling in ovarian cancer patients resulted in a disappointing clinical benefit. In this study, we found that EGFR could activate IL-6-STAT3 pathway in ovarian cancer cells. However, we also demonstrated that EGFR knockdown could increase STAT3 phosphorylation in HO8910 and OVCAR-3 ovarian cancer cells. Interestingly, we further demonstrated that the non-coding RNA miR-146b could simultaneously block both the EGFR and IL-6-STAT3 pathways. Finally, our data demonstrated that miR-146b overexpression resulted in a greater suppression of cell migration than STAT3 pathway inhibition alone.These results suggest a complex and heterogeneous role of EGFR in ovarian cancer. Combined blockade of EGFR and IL-6-STAT3 pathways by miR-146b might be a strategy for improving the clinical benefit of targeting the EGFR pathway in ovarian cancer patients in the future.

Preeclampsia (PE) is characterized by gestational hypertension and proteinuria, and is a leading cause of maternal death and perinatal morbidity globally. Although the exact cause of PE remains unclear, several studies have suggested a role for abnormal expression of multiple genes. The aim of the present study was to identify key genes and related pathways, and to screen for drugs that regulate these genes for potential PE therapy. The GSE60438 dataset was acquired from the Gene Expression Omnibus database to analyze differentially expressed genes (DEGs). By constructing a protein-protein interaction network and performing reverse transcription-quantitative PCR verification, proteasome 26S subunit, non-ATPase 14, prostaglandin E synthase 3 and ubiquinol-cytochrome c reductase core protein 2 were identified as key genes in PE. In addition, PE was found to be associated with ‘circadian rhythm’, ‘fatty acid metabolism’, ‘DNA damage response detection of DNA damage’, ‘regulation of DNA repair’ and ‘endothelial cell development’. Through connectivity map analysis of DEGs, furosemide and droperidol were suggested to be therapeutic drugs that may target the hub genes for PE treatment. Results analysis of GSEA were included in the discussion section of this article. In conclusion, the current study identified novel key genes associated with the onset of PE and potential drugs for PE treatment.

The failure of a massive influx of tumor-infiltrating T lymphocytes to eradicate tumor cells in the tumor microenvironment is mainly due to the dysfunction of T cells hyporesponsive to tumors. T-cell exhaustion and senescence induced by malignant tumors are two important dysfunctional states that coexist in cancer patients, hindering effective antitumor immunity and immunotherapy and sustaining the suppressive tumor microenvironment. Although exhausted and senescent T cells share a similar dysfunctional role in antitumor immunity, they are distinctly different in terms of generation, development, and metabolic and molecular regulation during tumor progression. Here, we discuss the unique phenotypic and functional characteristics of these two types of dysfunctional T cells and their roles in tumor development and progression. In addition, we further discuss the potential molecular and metabolic signaling pathways responsible for the control of T-cell exhaustion and senescence in the suppressive tumor microenvironment. Understanding these critical and fundamental features should facilitate rethinking the unresponsiveness to current immunotherapies in clinical patients and lead to further development of novel and effective strategies that target different types of dysfunctional T cells to enhance cancer immunotherapy.

RNA, one of the major building blocks of the cell, participates in many essential life processes. RNA stability is well-established to be closely related to various RNA modifications. To date, hundreds of different RNA modifications have been identified. N6-methyladenosine (m

Background The thymic microenvironment is mainly comprised of thymic epithelial cells, the cytokines, exosomes, surface molecules, and hormones from the cells, and plays a vital role in the development, differentiation, maturation and homeostasis of T lymphocytes. However, the thymus begins to degenerate as early as the second year of life and continues through aging in human beings, leading to a decreased output of naïve T cells, the limited TCR diversity and an expansion of monoclonal memory T cells in the periphery organs. These alternations will reduce the adaptive immune response to tumors and emerging infectious diseases, such as COVID-19, also it is easier to suffer from autoimmune diseases in older people. In the context of global aging, it is important to investigate and clarify the causes and mechanisms of thymus involution. Main body Epigenetics include histone modification, DNA methylation, non-coding RNA effects, and chromatin remodeling. In this review, we discuss how senescent thymic epithelial cells determine and control age-related thymic atrophy, how this process is altered by epigenetic modification. How the thymus adipose influences the dysfunctions of the thymic epithelial cells, and the prospects of targeting thymic epithelial cells for the treatment of thymus atrophy. Conclusion Epigenetic modifications are emerging as key regulators in governing the development and senescence of thymic epithelial cells. It is beneficial to re-establish effective thymopoiesis, identify the potential therapeutic strategy and rejuvenate the immune function in the elderly.

Multiple proinflammatory conditions, including chemotherapy, radiotherapy, transplant rejection, and microbial infections, have been identified to induce involution of the thymus. However, the underlying cellular and molecular mechanisms of these inflammatory conditions inducing apoptosis of thymic epithelial cells (TECs), the main components of the thymus, remain largely unknown. In the circulation, mature dendritic cells (mDCs), the predominant initiator of innate and adaptive immune response, can migrate into the thymus. Herein, we demonstrated that mDCs were able to directly inhibit TECs proliferation and induce their apoptosis by activating the Jagged1/Notch3 signaling pathway. Intrathymic injection of either mDCs or recombinant mouse Jagged1-human Fc fusion protein (rmJagged1-hFc) into mice resulted in acute atrophy of the thymus. Furthermore, DAPT, a γ-secretase inhibitor, reversed the effects induced by mDC or rmJagged1-hFc. These findings suggest that acute or aging-related thymus degeneration can be induced either by mass migration of circulating mDCs in a short period of time or by a few but constantly homing mDCs.

The subset of regulatory T (Treg) cells, with its specific transcription Foxp3, is a unique cell type for the maintenance of immune homeostasis by controlling effector T (Teff) cell responses. Although it is common that a defect in Treg cells with Treg/Teff disorder causes autoimmune diseases; however, the precise mechanisms are not thoroughly revealed. Here, we report that miR-34a could attenuate human and murine Foxp3 gene expression via targeting their 3′ untranslated regions (3′ UTR). The human miR-34a, increased in peripheral blood mononuclear cells (PBMCs) and CD4+ T cells from rheumatoid arthritis (RA) or systemic lupus erythematosus (SLE) patients, displayed a positive correlation with some serum markers of inflammation including rheumatoid factor (RF), anti-streptolysin antibody (ASO), erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) as well as Th17 signature gene RORγt, but inversely correlated with the mRNA expression levels of FOXP3. In addition, murine miR-34a levels were downregulated in TGF-β-induced Treg cells but upregulated in Th17 cells induced in vitro compared to activated CD4+ T cells. It has also been demonstrated that elevated miR-34a disrupting Treg/Th17 balance in vivo contributed to the progress of pathogenesis of collagen induced arthritis (CIA) mice. Furthermore, IL-6 and TNF-α were responsible for the upregulation of miR-34a and downregulation of Foxp3, which was reverted by the addition of NF-κB/p65 inhibitor BAY11-7082, thus indicating that NF-κB/p65 inhibited Foxp3 expression in an miR-34a-dependent manner. Finally, IL-6 or TNF-α-activated p65 could bind to the miR-34a promotor and enhance its activity, resulting in upregulation of its transcription. Taken together, we show that NF-κB activated by inflammatory cytokines, such as IL-6 and TNF-α, ameliorates Foxp3 levels via regulating miR-34a expression, which provides a new mechanistic and therapeutic insight into the ongoing of autoimmune diseases.

Epithelial ovarian cancer (EOC) is the most lethal gynecological malignancy. LIN28 homolog A (LIN28A) is a RNA-binding protein, which serves a fundamental role in cell development and pluripotency. Polo-like kinase 4 (PLK4) is a member of the polo-like kinase family, which primarily takes part in the mitotic regulation. Overexpression of LIN28A has been demonstrated in ovarian cancer; however, the expression of PLK4 and the correlation between the expression of LIN28A and PLK4 in EOC has not been discussed. In the present study, the mRNA and protein levels of LIN28A and PLK4 were evaluated by reverse transcription-quantitative polymerase chain reaction and immunohistochemistry in ovarian tissues of patients. Results demonstrated significantly increased expression in EOC compared with benign epithelial ovarian tumors. High expression of LIN28A and PLK4 was detected at the advanced pathological stage. Furthermore, PLK4 expression was positively correlated with LIN28A (r=0.555; P=0.039). The median survival analysis of patients with EOC with LIN28A and PLK4 double positive expression was 14 months, compared with 30 months in single positive and 60 months in double negative patients by Kaplan-Meier analysis (P

Pyroptosis is a recently identified mechanism of programmed cell death related to Caspase-1 that triggers a series of inflammatory reactions by releasing several proinflammatory factors such as IL-1β and IL-18. The process is characterised by the rupture of cell membranes and the release of cell contents through the mediation of gasdermin (GSDM) proteins. GSDMD is an important member of the GSDM family and plays a critical role in the two pathways of pyroptosis. Diabetic nephropathy (DN) is a microvascular complication of diabetes and a major cause of end-stage renal disease. Recently, it was revealed that GSDMD-mediated pyroptosis plays an important role in the occurrence and development of DN. In this review, we focus on two types of kidney cells, tubular epithelial cells and renal podocytes, to illustrate the mechanism of pyroptosis in DN and provide new ideas for the prevention, early diagnosis and molecular therapy of DN.

Much attention has been paid to a newly discovered subset of memory T (TM) cells-stem cell-like memory T (TSCM) cells for their high self-renewal ability, multi-differentiation potential and long-term effector function in adoptive therapy against tumors. Despite their application in cancer therapy, an excess of TSCM cells also contributes to the persistence of autoimmune diseases for their immune memory and HIV infection as a long-lived HIV reservoir. Signaling pathways Wnt, AMPK/mTOR and NF-κB are key determinants for TM cell generation, maintenance and proinflammatory effect. In this review, we focus on the phenotypic and functional characteristics of TSCM cells and discuss their role in autoimmune diseases and HIV-1 chronic infection. Also, we explore the potential mechanism and signaling pathways involved in immune memory and look into the future therapy strategies of targeting long-lived TM cells to suppress pathogenic immune memory.

Regulatory factor X-5 (RFX5) represents a key transcription regulator of MHCII gene expression in the immune system. This study aims to explore the molecular mechanisms and biological significance of RFX5. Firstly, by analyzing ENCODE chromatin immunoprecipitation (ChIP)-seq in HepG2 and TCGA RNA-seq data, we discovered lysine-specific demethylase 4A (KDM4A), also named JMJD2A, to be a major downstream target gene of RFX5. Moreover, RFX5 was verified to bind directly to the KDM4A’s promoter region and sequentially promoted its transcription determined by the ChIP-PCR assay and luciferase assay. In addition, RFX5-dependent regulation of KDM4A was demonstrated in HCC. Compared with adjacent non-tumor tissues, the expression levels of KDM4A were significantly raised in HCC tumor tissues. Notably, elevated levels of KDM4A were strongly correlated with HCC patient prognosis. Functionally, KDM4A overexpression largely rescued the growth inhibitory effects of RFX5 deletion, highlighting KDM4A as a downstream effector of RFX5. Mechanistically, the RFX5-KDM4A pathway promoted the progression of the cell cycle from G0/G1 to S phase and was protective against cell apoptosis through regulation of p53 and its downstream genes in HCC. In conclusion, RFX5 could promote HCC progression via transcriptionally activating KDM4A expression.

Metabolic disorder is one of the hallmarks of cancers, and reprogramming of metabolism is becoming a novel strategy for cancer treatment. Citrate is a key metabolite and critical metabolic regulator linking glycolysis and lipid metabolism in cellular energy homeostasis. Here it is reported that citrate treatment (both sodium citrate and citric acid) significantly suppresses tumor cell proliferation and growth in various tumor types. Mechanistically, citrate promotes excessive lipid biosynthesis and induces disruption of lipid metabolism in tumor cells, resulting in tumor cell senescence and growth inhibition. Furthermore, ATM‐associated DNA damage response cooperates with MAPK and mTOR signaling pathways to control citrate‐induced tumor cell growth arrest and senescence. In vivo studies further demonstrate that citrate administration dramatically inhibits tumor growth and progression in a colon cancer xenograft model. Importantly, citrate administration combined with the conventional chemotherapy drugs exhibits synergistic antitumor effects in vivo in the colon cancer models. These results clearly indicate that citrate can reprogram lipid metabolism and cell fate in cancer cells, and targeting citrate can be a promising therapeutic strategy for tumor treatment., Reprogramming of metabolism is a novel strategy for cancer treatment. Extracellular citrate administration in tumor cells induces activation of the DNA damage response, MAPK, and mTOR signaling pathways. The activated cascades further induce excessive lipid biosynthesis and provoke senescence to impair tumor cell growth in vitro and in vivo. Targeting citrate can be a promising therapeutic strategy for tumor treatment.

DNA microarray and high-throughput sequencing have been widely used to identify the differentially expressed genes (DEGs) in systemic lupus erythematosus (SLE). However, the big data from gene microarrays are also challenging to work with in terms of analysis and processing. The presents study combined data from the microarray expression profile (GSE65391) and bioinformatics analysis to identify the key genes and cellular pathways in SLE. Gene ontology (GO) and cellular pathway enrichment analyses of DEGs were performed to investigate significantly enriched pathways. A protein‑protein interaction network was constructed to determine the key genes in the occurrence and development of SLE. A total of 310 DEGs were identified in SLE, including 193 upregulated genes and 117 downregulated genes. GO analysis revealed that the most significant biological process of DEGs was immune system process. Kyoto Encyclopedia of Genes and Genome pathway analysis showed that these DEGs were enriched in signaling pathways associated with the immune system, including the RIG‑I‑like receptor signaling pathway, intestinal immune network for IgA production, antigen processing and presentation and the toll‑like receptor signaling pathway. The current study screened the top 10 genes with higher degrees as hub genes, which included 2'‑5'‑oligoadenylate synthetase 1, MX dynamin like GTPase 2, interferon induced protein with tetratricopeptide repeats 1, interferon regulatory factor 7, interferon induced with helicase C domain 1, signal transducer and activator of transcription 1, ISG15 ubiquitin‑like modifier, DExD/H‑box helicase 58, interferon induced protein with tetratricopeptide repeats 3 and 2'‑5'‑oligoadenylate synthetase 2. Module analysis revealed that these hub genes were also involved in the RIG‑I‑like receptor signaling, cytosolic DNA‑sensing, toll‑like receptor signaling and ribosome biogenesis pathways. In addition, these hub genes, from different probe sets, exhibited significant co‑expressed tendency in multi‑experiment microarray datasets (P

// Hui Wang 1 , Wenrong Xu 2 , Qixiang Shao 1 and Qing Ding 1 1 Jiangsu Key Laboratory of Medical Science and Laboratory Medicine, Department of Immunology, School of Medicine, Jiangsu University, Zhenjiang, Jiangsu, P.R. China 2 Key Laboratory of Laboratory Medicine of Jiangsu Province, School of Medicine, Jiangsu University, Zhenjiang, Jiangsu, P.R. China Correspondence to: Qixiang Shao, email: shao_qx@ujs.edu.cn Qing Ding, email: qing_ding69@163.com Keywords: microRNA-21, interleukin-10, regulatory B cells, experimental autoimmune encephalomyelitis Received: June 29, 2017 Accepted: September 18, 2017 Published: October 06, 2017 ABSTRACT IL-10-producing regulatory B (IL-10 + Breg) cells promote tolerance in autoimmune diseases and transplantation. However, it remains unclear whether microRNAs are involved in the development of IL-10 + Breg cells. Here, we found that microRNA-21 (miR-21) acts as an upstream regulator of IL-10 by targeting the 3' untranslated region of IL-10 mRNA. We also demonstrated that IL-10 + Breg cells exhibit lower miR-21 expression than non-Breg cells and that miR-21 acts as a potent negative regulator of the differentiation of IL-10 + Breg cells. Accordingly, specific inhibition of miR-21 using antisense oligonucleotides markedly promoted B cell IL-10 expression. Thus, IL-10 is a direct target of miR-21. Moreover, silencing of miR-21 significantly alleviated the severity of experimental autoimmune encephalomyelitis (EAE), and this change was associated with an increase in the number of IL-10 + Breg cells. Finally, we demonstrated that miR-21-silenced B cells exert their suppressive activity through effector T cells in an IL-10-dependent manner. Thus, we characterized a B cell-intrinsic microRNA pathway that inhibits the differentiation of IL-10 + Breg cells and promotes autoimmunity. miR-21 silencing therefore represents a new therapeutic strategy for the treatment of autoimmune diseases.

Laboratory exercise is helpful for medical students to understand the basic principles of molecular biology and to learn about the practical applications of molecular biology. We have designed a lab course on molecular biology about the determination of single nucleotide polymorphism (SNP) in human REV3 gene, the product of which is a subunit of DNA polymerase ζ and SNPs in this gene are associated with altered susceptibility to cancer. This newly designed experiment is composed of three parts, including genomic DNA extraction, gene amplification by PCR, and genotyping by RFLP. By combining these activities, the students are not only able to learn a series of biotechniques in molecular biology, but also acquire the ability to link the learned knowledge with practical applications. This comprehensive experiment will help the medical students improve the conceptual understanding of SNP and the technical understanding of SNP detection. © 2017 by The International Union of Biochemistry and Molecular Biology, 45(4):299-304, 2017.

Forkhead box P3 (FOXP3), which is a transcription factor, has a primary role in the development and function of regulatory T cells, and thus contributes to homeostasis of the immune system. A previous study generated a cell-permeable fusion protein of mouse FOXP3 conjugated to a protein transduction domain (PTD-mFOXP3) that successfully blocked differentiation of type 17 T helper cells in vitro and alleviated experimental arthritis in mice. In the present study, the role of PTD-mFOXP3 in type 1 T helper (Th1) cell-mediated immunity was investigated and the possible mechanisms for its effects were explored. Under Th1 polarization conditions, cluster of differentiation 4+ T cells were treated with PTD-mFOXP3 and analyzed by flow cytometry in vitro, which revealed that PTD-mFOXP3 blocked Th1 differentiation in vitro. Mice models of delayed type hypersensitivity (DTH) reactions were generated by subcutaneous sensitization and challenge with ovalbumin (OVA) to the ears of mice. PTD-mFOXP3, which was administered via local subcutaneous injection, significantly reduced DTH-induced inflammation, including ear swelling (ear swelling, P

Epidemiological studies have shown that fat rich western diet contributes to the high incidence of inflammatory bowel disease (IBD). Moreover, accumulated data indicated that fat dietary factor might promote the change of the composition and metabolism in commensal flora. But, the exact mechanisms for fatty diet in gut inflammation are not well demonstrated. In this study, we found that high fat diet (HFD) promoted inflammation and exacerbated the disease severity of dextran sulfate sodium (DSS) induced colitis in mice. Compared with low fat diet (LFD)/DSS mice, shorter colon length, more epithelial loss and crypt destruction and more Gr-1+ myeloid inflammatory cells infiltration in colons were observed in HFD/DSS cohorts. Interestingly, such HFD mediated inflammation accompanied with the dys-regulation of hematopoiesis, and more hematopoiesis stem and progenitor cells were detected in colon and spleen. We further analyzed the effects of HFD and DSS treatment on mucosal DC subsets, and found that DSS treatment in LFD mice mainly dramatically increased the percentage of CD11c+CD103-CD11b+ DCs in lamina propria (LP). While, in HFD/DSS mice, HFD pre-treatment not only increased the percentage of CD11c+CD103-CD11b+ DCs, but also decreased CD11c+CD103+CD11b+ in both LP and mesenteric lymph nodes (MLN) in mice with colitis. This disequilibrium of mucosal dendritic cells in HFD/DSS mice may depend on the reduced levels of buytrate and retinoic acid. Thus, this study declared the effects of HFD on gut microenviroment, and further indicated its potential role in the development of DSS induced colitis.

Construction of a random ssDNA sublibrary is an important step of the aptamer screening process. The available construction methods include asymmetric PCR, biotin-streptavidin separation, and lambda exonuclease digestions, in which PCR amplification is a key step. The main drawback of PCR amplification is overamplification increasing nonspecific hybridization among different products and by-products, which may cause the loss of potential high-quality aptamers, inefficient screening, and even screening failure. Cycle number optimization in PCR amplification is the main way to avoid overamplification but does not fundamentally eliminate the nonspecific hybridization, and the decreased cycle number may lead to insufficient product amounts. Here, we developed a new method, "asymmetric emulsion PCR," which could overcome the shortcomings of conventional PCR. In asymmetric emulsion PCR, different templates were separated by emulsion particles, allowing single-molecule PCR, in which each template was separately amplified, and the nonspecific hybridization was avoided. Overamplification or formation of by-products was not observed. The method is so simple that direct amplification of 40 or more cycles can provide a high-quality ssDNA library. Therefore, the asymmetric emulsion PCR would improve the screening efficiency of systematic evolution of ligands by exponential enrichment.

Self-renewal and differentiation in germline stem cells (GSCs) are tightly regulated by the stem cell niche and via multiple approaches. In our previous study, we screened the novel GSC regulatory gene Srlp in Drosophila testes. However, the underlying mechanistic links between Srlp and the stem cell niche remain largely undetermined. Here, using genetic manipulation of the Drosophila model, we systematically analyze the function and mechanism of Srlp in vivo and in vitro. In Drosophila, Srlp is an essential gene that regulates the self-renewal and differentiation of GSCs in the testis. In the in vitro assay, Srlp is found to control the proliferation ability and cell death in S2 cells, which is consistent with the phenotype observed in Drosophila testis. Furthermore, results of the liquid chromatography-tandem mass spectrometry (LC-MS/MS) reveal that RpL6 binds to Srlp. Srlp also regulates the expression of spliceosome and ribosome subunits and controls spliceosome and ribosome function via RpL6 signals. Collectively, our findings uncover the genetic causes and molecular mechanisms underlying the stem cell niche. This study provides new insights for elucidating the pathogenic mechanism of male sterility and the formation of testicular germ cell tumor.

Germ cell maturation is essential for spermatogenesis and testis homeostasis. ATP synthase serves significant roles in energy storage in germ cell survival and is catalyzed by alterations in the mitochondrial membrane proton concentration. The intrinsic cellular mechanisms governing stem cell maturation remain largely unknown. In the present study, in vivo RNA interference (RNAi) screening of major ATP synthase subunits was performed, and the function of ATP synthase for male fertility and spermatogenesis in Drosophila was explored. A Upstream Activation Sequence/Gal4 transcription factor system was used to knock down gene expression in specific cell types, and immunofluorescence staining was conducted to assess the roles of ATP synthase subunits in Drosophila testes. It was identified that knockdown of ATP synthase resulted in male infertility and abnormal spermatogenesis in Drosophila testes. In addition, knockdown of the ATP synthase β subunit in germ cells resulted in defects in male infertility and germ cell maturation, while the hub and cyst cell populations were maintained. Other major ATP synthase subunits were also examined and similar phenotypes in Drosophila testes were identified. Taken together, the data from the present study revealed that ATP synthase serves important roles for male fertility during spermatogenesis by regulating germ cell maturation in Drosophila testes.