Tissue-specific cancer stem/progenitor cells:Therapeutic implications

Surgical resection,chemotherapy,and radiation are the standard therapeutic modalities for treating cancer.These approaches are intended to target the more mature and rapidly dividing cancer cells.However,they spare the relatively quiescent and intrinsically resistant cancer stem cells(CSCs)subpopulation residing within the tumor tissue.Thus,a temporary eradication is achieved and the tumor bulk tends to revert supported by CSCs'resistant features.Based on their unique expression profile,the identification,isolation,and selective targeting of CSCs hold great promise for challenging treatment failure and reducing the risk of cancer recurrence.Yet,targeting CSCs is limited mainly by the irrelevance of the utilized cancer models.A new era of targeted and personalized anti-cancer therapies has been developed with cancer patient-derived organoids(PDOs)as a tool for establishing pre-clinical tumor models.Herein,we discuss the updated and presently available tissue-specific CSC markers in five highly occurring solid tumors.Additionally,we highlight the advantage and relevance of the threedimensional PDOs culture model as a platform for modeling cancer,evaluating the efficacy of CSC-based therapeutics,and predicting drug response in cancer patients.

Tissue-specific differential expression of novel genes and long intergenic non-coding RNAs in humans with extreme response to evoked endotoxemia

Objective Cytokine responses to activation of innate immunity differ between individuals,yet the genomic and tissue-specific transcriptomic determinants of inflammatory responsiveness are not well understood. We hypothesized that tissue-specific mRNA and long intergenic non-coding RNA (lincRNA) induction differs between individuals with divergent evoked inflammatory responses.

Development of a Novel Tissue-Specific Method to Detect Cytokeratin 20-Positive Circulating Tumor Cells in Metastatic Colorectal Cancer

Introduction:Although many studies have shown the vast potential of circulating tumor cells(CTCs)detection in cancer diagnosis and prognosis,our understanding of their clinical significance is still far from complete.A major obstacle arises from the lack of well-established tumor or tissue-specific markers to detect CTCs by immunocytochemical staining after immunomagnetic enrichment(IE).Methods:We have established the utility of cytokeratin 20(CK20),a gastrointestinal tract specific marker,for the specific detection and identification of colorectal cancer(CRC)CTCs.This breakthrough was successfully validated in spike-in experiments using CRC cell line models followed by a pilot study which recruited 32 metastatic CRC patients,25 benign colorectal diseases patients and 27 normal subjects.Results:CK20-positive CTCs were detected in 90%metastatic CRC patients but not in benign colorectal diseases patients and normal subjects using this refined assay.Conclusions:These impressive results have laid the foundation for further development of CK20-positive CTCs as a promising marker in diagnosis,prognostication and treatment monitoring of metastatic CRC.

Matrix from urine stem cells boosts tissue-specific stem cell mediated functional cartilage reconstruction

Articular cartilage has a limited capacity to self-heal once damaged.Tissue-specific stem cells are a solution for cartilage regeneration;however,ex vivo expansion resulting in cell senescence remains a challenge as a large quantity of high-quality tissue-specific stem cells are needed for cartilage regeneration.Our previous report demonstrated that decellularized extracellular matrix(dECM)deposited by human synovium-derived stem cells(SDSCs),adipose-derived stem cells(ADSCs),urine-derived stem cells(UDSCs),or dermal fibroblasts(DFs)provided an ex vivo solution to rejuvenate human SDSCs in proliferation and chondrogenic potential,particularly for dECM deposited by UDSCs.To make the cell-derived dECM(C-dECM)approach applicable clinically,in this study,we evaluated ex vivo rejuvenation of rabbit infrapatellar fat pad-derived stem cells(IPFSCs),an easily accessible alternative for SDSCs,by the abovementioned C-dECMs,in vivo application for functional cartilage repair in a rabbit osteochondral defect model,and potential cellular and molecular mechanisms underlying this rejuvenation.We found that C-dECM rejuvenation promoted rabbit IPFSCs’cartilage engineering and functional regeneration in both ex vivo and in vivo models,particularly for the dECM deposited by UDSCs,which was further confirmed by proteomics data.RNA-Seq analysis indicated that both mesenchymal-epithelial transition(MET)and inflammation-mediated macrophage activation and polarization are potentially involved in the C-dECM-mediated promotion of IPFSCs’chondrogenic capacity,which needs further investigation.

Genome-wide Transcription Factor Gene Prediction and their Expressional Tissue-Specificities in Maize

Transcription factors （TFs） are important regulators of gene expression. To better understand TFencoding genes in maize （Zea mays L.）, a genome-wide TF prediction was performed using the updated B73 reference genome. A total of 2 298 TF genes were identified, which can be classified into 56 families. The largest family, known as the MYB superfamily, comprises 322 MYB and MYB-related TF genes. The expression patterns of 2014 （87.64%） TF genes were examined using RNA-seq data, which resulted in the identification of a subset of TFs that are specifically expressed in particular tissues （including root, shoot, leaf, ear, tassel and kernel）. Similarly, 98 kernel-specific TF genes were further analyzed, and it was observed that 29 of the kernel-specific genes were preferentially expressed in the early kernel developmental stage, while 69 of the genes were expressed in the late kernel developmental stage. Identification of these TFs, particularly the tissue-specific ones, provides important information for the understanding of development and transcriptional regulation of maize.

Rice tissue-specific promoters and conditiondependent promoters for effective translational application

Rice （Oryza sativa） is one of the most important staple food crops for more than half of the world＇s population. The demand is increasing for food security because of population growth and environmental challenges triggered by climate changes. This scenario has led to more interest in developing crops with greater productivity and sustainability. The process of genetic transformation, a major tool for crop improvement, utilizes promoters as one of its key elements. Those promoters are generally divided into three types： constitutive, spatiotemporal, and condition-dependent. Tran- scriptional control of a constitutive promoter often leads to reduced plant growth, due to a negative effect of accumu- lated molecules during cellular functions or energy consump- tion. To maximize the effect of a transgene on transgenic plants, it is better to use condition-dependent or tissue- specific promoters. However, until now, those types have not been as widely applied in crop biotechnology. In this review, we introduce and discuss four groups of tissue-specific promoters （5o promoters in total） and six groups of condition-dependent promoters （27 promoters）. These pro- moters can be utilized to fine-tune desirable agronomic traits and develop crops with tolerance to various stresses, enhanced nutritional value, and advanced productivity.

Transgenic Bt cotton driven by the green tissue-specific promoter shows strong toxicity to lepidopteran pests and lower Bt toxin accumulation in seeds

A promoter of the PNZIP （Pharbitis nil leucine zipper） gene （1.459 kb） was cloned from Pharbitis nil and fused to the GUS（^-glucuronidase） and Bacillus thuringiensis endotoxin （Cry9C） genes. Several transgenic PNZIP：：GUS and PNZIP：：Cry9C cotton lines were developed by Agrobacterium-mediated transformation. Strong GUS staining was detected in the green tissues of the transgenic PNZIP：：GUS cotton plants. In contrast, GUS staining in the reproductive structures such as petals, anther, and immature seeds of PNZIP：：GUS cotton was very faint. Two transgenic PNZIP：：Cry9C lines and one trans- genic cauliflower mosaic virus （CaMV） 35S：：Cry9C line were selected for enzyme-linked immunosorbent assay （ELISA） and insect bioassays. Expression of the Cry9C protein in the 35S：：Cry9C line maintained a high level in most tissues ranging from 24.6 to 45.5 ~tg g-I fresh weight. In green tissues such as the leaves, boll rinds, and bracts of the PNZIP：：Cry9C line, the Cry9C protein accumulated up to 50.2, 39.7, and 48.3 jag g-a fresh weight respectively. In contrast, seeds of the PNZIP：：Cry9C line （PZ1.3） accumulated only 0.26 ~ag g-~ fresh weight of the Cry9C protein, which was 100 times lower than that recorded for the seeds of the CaMV 35S：：Cry9C line. The insect bioassay showed that the transgenic PNZIP：：Cry9C cotton plant exhibited strong resistance to both the cotton bollworm and the pink bollworm. The PNZIP promoter could effectively drive Bt toxin ex- pression in green tissues of cotton and lower accumulated levels of the Bt protein in seeds. These features should allay public concerns about the safety of transgenic foods. We propose the future utility of PNZIP as an economical, environmentally friendly promoter in cotton biotechnology.

Experimental Study Enhancing the Chemosensitivityof Multiple Myeloma to Melphalan by Using a Tissue-Specific APE1-Silencing RNA Expression Vector

Introduction:Multiple myeloma (MM) is a geriatric disease with onset at an average age of approximately 61 years.With the aging of the population,the incidence rate of MM is climbing.In the United States,the annual incidence rate of MM is 2-5/100,000.Multiple myeloma accounts for approximately 1% of all tumor eases and slightly 】10% of cases with hematologic malignancy.Although an

A Systematic Phenotypic Screen of F-box Genes Through a Tissue-specific RNAi-based Approach in Drosophila

F-box proteins are components of the SCF （SkpA-Cullin 1-F-box） E3 ligase complexes, acting as the specificity-determinants in targeting substrate proteins for ubiquitination and degradation. In humans, at least 22 out of 75 F-box proteins have experimentally documented substrates, whereas in Drosophila 12 F-box proteins have been characterized with known substrates. To systematically investigate the genetic and molecular functions of F-box proteins in Drosophila, we performed a survey of the literature and databases. We identified 45 Drosophila genes that encode proteins containing at least one F-box domain. We collected publically available RNAi lines against these genes and used them in a tissue-specific RNAi-based phenotypic screen. Here, we present our systematic phenotypic dataset from the eye, the wing and the notum. This dataset is the first of its kind and represents a useful resource for future studies of the molecular and genetic functions of F-box genes in Drosophila. Our results show that, as expected, F-box genes in Drosophila have regulatory roles in a diverse array of processes including cell proliferation, cell growth, signal transduction, and cellular and animal survival.

Proteomic survey towards the tissue-specific proteins of mouse mitochondria

Mitochondrion plays the key functions in mammalian cells. It is believed that mitochondrion exerts the common biologic functions in many tissues, but also performs some specific functions correspondent with tissues where it is localized. To identify the tissue-specific mitochondrial proteins, we carried out a systematic survey towards mitochondrial proteins in the tissues of C57BL/6J mouse, such as liver, kidney and heart. The mitochondrial proteins were separated by 2DE and identified by MALDI-TOF/TOF MS. Total of 87 unique proteins were identified as the tissue-specific ones, and some representatives were further verified through ICPL quantification and Western blot. Because these issue-specific proteins are coded from nuclear genes, real-time PCR was employed to examine the mRNA status of six typical genes found in the tissues.With combining of the expression data and the co-localization images obtained from confocal microscope, we came to the conclusion that the tissue-specifically mitochondrial proteins were widely distributed among the mouse tissues. Our investigation, therefore, indeed provides a solid base to further explore the biological significance of the mitochondrial proteins with tissue-orientation.

Tissue-specific Hi-C analyses of rice, foxtail millet and maize suggest non-canonical function of plant chromatin domains

Chromatins are not randomly packaged in the nucleus and their organization plays important roles in transcription regulation,which is best studied in the mammalian models.Using in situ Hi-C,we have compared the 3D chromatin architectures of rice mesophyll and endosperm,foxtail millet bundle sheath and mesophyll,and maize bundle sheath,mesophyll and endosperm tissues.We found that their global A/B compartment partitions are stable across tissues,while local A/B compartment has tissue-specific dynamic associated with differential gene expression.Plant domains are largely stable across tissues,while new domain border formations are often associated with transcriptional activation in the region.Genes inside plant domains are not conserved across species,and lack significant co-expression behavior unlike those in mammalian TADs.Although we only observed chromatin loops between gene islands in the large genomes,the maize loop gene pairs’syntenic orthologs have shorter physical distances in small genome monocots,suggesting that loops instead of domains might have conserved biological function.Our study showed that plants’chromatin features might not have conserved biological functions as the mammalian ones.

Simultaneous inference of phenotype-associated genes and relevant tissues from GWAS data via Bayesian integration of multiple tissue-specific gene networks

Although genome-wide association studies （GWAS） have successfully identified thousands of genomic loci associated with hun- dreds of complex traits in the past decade, the debate about such problems as missing heritabiUty and weak interpretability has been appealing for effective computational methods to facilitate the advanced analysis of the vast volume of existing and antici- pated genetic data. Towards this goal, gene-tevel integrative GWAS analysis with the assumption that genes associated with a phenotype tend to be enriched in biological gene sets or gene networks has recently attracted much attention, due to such advan- tages as straightforward interpretation, tess multiple testing burdens, and robustness across studies. However, existing methods in this category usually exploit non-tissue-specific gene networks and thus lack the ability to utilize informative tissue-specific characteristics. To overcome this limitation, we proposed a Bayesian approach called SIGNET （Simultaneously Inference of GeNEs and Tissues） to integrate GWAS data and multiple tissue-specific gene networks for the simultaneous inference of phenotype- associated genes and relevant tissues. Through extensive simulation studies, we showed the effectiveness of our method in find- ing both associated genes and relevant tissues for a phenotype. In applications to real GWAS data of 14 complex phenotypes, we demonstrated the power of our method in both deciphering genetic basis and discovering biological insights of a phenotype. With this understanding, we expect to see SIGNET as a valuable tool for integrative GWAS analysis, thereby boosting the preven- tion, diagnosis, and treatment of human inherited diseases and eventually facilitating precision medicine.

Constructing a highly bioactive tendon-regenerative scaffold by surface modification of tissue-specific stem cell-derived extracellular matrix

Developing highly bioactive scaffold materials to promote stem cell migration,proliferation and tissue-specific differentiation is a crucial requirement in current tissue engineering and regenerative medicine.Our previous work has demonstrated that the decellularized tendon slices(DTSs)are able to promote stem cell proliferation and tenogenic differentiation in vitro and show certain pro-regenerative capacity for rotator cuff tendon regeneration in vivo.In this study,we present a strategy to further improve the bioactivity of the DTSs for constructing a novel highly bioactive tendon-regenerative scaffold by surface modification of tendon-specific stem cell-derived extracellular matrix(tECM),which is expected to greatly enhance the capacity of scaffold material in regulating stem cell behavior,including migration,proliferation and tenogenic differentiation.We prove that the modification of tECM could change the highly aligned surface topographical cues of the DTSs,retain the surface stiffness of the DTSs and significantly increase the content of multiple ECM components in the tECM-DTSs.As a result,the tECM-DTSs dramatically enhance the migration,proliferation as well as tenogenic differentiation of rat bone marrow-derived stem cells compared with the DTSs.Collectively,this strategy would provide a new way for constructing ECMbased biomaterials with enhanced bioactivity for in situ tendon regeneration applications.

Chicken chromatin accessibility atlas accelerates epigenetic annotation of birds and gene fine-mapping associated with growth traits

The development of epigenetic maps, such as the ENCODE project in humans, provides resources for gene regulation studies and a reference for research of disease-related regulatory elements. However,epigenetic information, such as a bird-specific chromatin accessibility atlas, is currently lacking for the thousands of bird species currently described.The major genomic difference between birds and mammals is their shorter introns and intergenic distances, which seriously hinders the use of humans and mice as a reference for studying the function of important regulatory regions in birds. In this study, using chicken as a model bird species, we systematically compiled a chicken chromatin accessibility atlas using 53 Assay of Transposase Accessible Chromatin sequencing(ATAC-seq)samples across 11 tissues. An average of 50 ?796open chromatin regions were identified per sample,cumulatively accounting for 20.36% of the chicken genome. Tissue specificity was largely reflected by differences in intergenic and intronic peaks, with specific functional regulation achieved by two mechanisms: recruitment of several sequence-specific transcription factors and direct regulation of adjacent functional genes. By integrating data from genome-wide association studies, our results suggest that chicken body weight is driven by different regulatory variants active in growth-relevant tissues. We propose CAB39L(active in the duodenum), RCBTB1(muscle and liver), and novel long non-coding RNA ENSGALG00000053256(bone) as candidate genes regulating chicken body weight. Overall, this study demonstrates the value of epigenetic data in fine-mapping functional variants and provides a compendium of resources for further research on the epigenetics and evolution of birds and mammals.

Isozyme Analysis of Spotted Halibut Verasper variegatus Temminck et Schlegel

To investigate the tissue-specificities of isozymes and the genetic structureof wild spotted halibut ( Verasper variegatus) population, horizontal starch gel electrophoresiswas performed on 45 individuals collected in part of the Yellow Sea. The performances of 17 isozymesin 8 kinds of tissues or organs were screened preliminarily in a TC-7.0 buffer system. The resultsshowed that the screened isozymes displayed remarkable tissue-specificities. Finally, 14 enzymes(AAT, ADH, EST, OPI, G3PDH, IDHP, LAP, LDH, MDH, MPI, PGDH, PGM, SDH and SOD) and 4 kinds of tissues(eye, skeleton muscle, liver and heart) were selected for genetic analysis. Fourteen isozymes areencoded by 20 loci, and 9 of them are polymorphic. The polymorphic loci are AAT-1~*, GPI-2~*,G3PDH~*, IDHP-1~*, LDH~*, MPI~*, PGM-1~*. PGM-2~* and SDH~*, and the proportion of polymorphic lociis 0.4500 (P_(0.99)) ? The mean values of observed and expected heterozygosities are 0.0278and 0.0265, respectively and the average effective number of alleles is 1.0675.

Why Mesenchymal Stem/Progenitor Cell Heterogeneity in Specific Environments? <br/>—Implications for Tissue Engineering Applications Following Injury or Degeneration of Connective Tissues

Mesenchymal stem/progenitor cells (MSC/MPC) from a variety of tissue sources (bone marrow, adipose tissue, fat pads, synovial membranes, synovial fluid, skin, muscle and periosteal tissue) have been widely applied for tissue engineering applications to generate replacements for injured or degenerated tissues. Alternatively, they have also been injected as free cells in an attempt to facilitate in vivo repair. Nearly all studies reported have used mixed cell populations of MSC/MPC, usually defined by cell surface phenotypes and/or functional ability to differentiate towards multiple cell lineages. Using more detailed cell surface phenotyping and limiting dilution approaches to isolate individual MSC/MPC clones have indicated that such mixed cell populations are very heterogeneous. In addition subsets of cells from different sources may have epigenetic modifications. While it is clear that MSC/MPC cells exhibit heterogeneity, the question of why this is the case has not been well addressed. This review will address some of these issues, as well as provide some insights into the implications when using such diverse cells for tissue engineering applications.

Genotypic Variation in Spatial Distribution of Fe in Rice Grains in Relation to Phytic Acid Content and Ferritin Gene Expression

Rice varieties having high Fe concentration in the endospermic region can be used as a good source for Fe deficit population.In this study,303 Oryza sativa varieties and 1 Oryza rufipogon accession were assessed for spatial Fe accumulation in grains by Prussian blue staining method.Spatial ferritin protein distribution in grains was visualized by immunohistochemistry,and ferritin expression was assessed in selected rice varieties using semi-quantitative reverse transcription PCR.Three popular rice varieties,namely Sarjoo 52,Madhukar and Jalmagna,and the O.rufipogon variety showed Fe in all the regions of grains,and the highest Fe concentration was observed in the embryo region.Some high-yielding varieties like Swarna,Swarna Sub 1,CSR13 and NDRR359 had lower Fe concentration in the embryo region.The highest Fe concentration was detected in O.rufipogon(49.8μg/g),followed by Sarjoo 52(26.1μg/g)and Madhukar(25.7μg/g).Phytic acid concentration was the minimum in O.rufipogon(5.75 mg/g)followed by Sarjoo 52(5.83 mg/g).Western blot and semi-quantitative reverse transcription PCR showed higher expression of ferritin gene in O.rufipogon,Sarjoo 52 and Madhukar.In conclusion,O.rufipogon and Sarjoo 52 had higher Fe concentration in the embryo regions as well as endosperm and aleurone layer,whereas the other varieties had lower Fe concentration in the endosperm.Sarjoo 52 could be used as a donor in the rice breeding program for the generation of new varieties with elevated grain Fe concentration.

Construction of retroviral vectors to induce a strong expression of human interferon-β gene in human hepatoma cells

Establishing the hepatoma cell-specific expression of human interferon gene mediated by retroviral vectors. Methods: Human interferon-β complementary DNA (IFN-β cDNA) was inserted into polylinker site of pMNSM retroviral vector to construct recombinant retroviral vector pMNSIFNB, where the transcription of IFN-β gene was driven by SV40 early region promoter, and MNAIFNB, where the transcription of IFN-β gene was driven by SV40 early region promoter regulated by α-fetoprotein enhancer. The retroviral constructs were respectively introduced into PA317 amphotropic packaging cells by means of lipofectamine mediated gene transfer procedure. The plasmids transfection efficiency was among (4-25)ｘ103 colonies/μg DNA/106 PA317 cells. The retrovirus infection efficiency was among (4. 5-500)ｘ104 Colony Forming Units (CFU)/ml. The recombinant retroviruses were used to infect human hepatoma cells, renal cell carcinoma cells and melanoma cell lines in the presence of 4 μg/ml polybrene. Results: Dot hybridization of total RNA from the neomycin resistant colonies and interferon expression assay indicated that human α-fetoprotein enhancer induced efficient and specific transcription and expression of IFN-β gene driven by the promoter of different origin in human hepatoma cells by which α-fetoprotein was highly produced. Conclusion: Cis-active element of α-fetoprotein gene can drive IFN-β gene specifically expressed in human hepatoma cells, which presents some valuable materials for the hepatoma-specific immune gene therapy.

Transcription Activity of Ectogenic Human Carcinoembryonic Antigen Promoter in Lung Adenocarcinoma Cells A549

The transcription activity of ectogenic human carcinoembryonic antigen （CEA） promoter in lung adenocarcinoma cells A549 was investigated for the further gene-targeting therapy. The reporter gene green fluorescent protein （GFP） driven by CEA promoter and human cytomegalovirus （CMV） promoter were relatively constructed and named plasmid pCEA-EGFP and pCMV-GFP respectively. The intensity of fluorescence was detected by fluorescence microscope and flow cytometry analysis after the pCEA-GFP and pSNAV-GFP plasmids were transfected into A549 cells through liposome respectively. The results showed （4,08±0.63） % of the A549 cells transfected with pCEA-AFP plasmid expressed, significantly lower than that of the A549 cells transfected with pCMV-GFP [（43.27±3.54） %]. It was suggested that ectogenic human CEA promoter in lung adenocarcinoma cells A549 was weakly expressed. The distinct specificity of CEA promoter in CEA high expression cells was regarded as a tool in selective gene therapy, but the transcription activity of ectogenic human CEA promoter was needed to increase in the future.

Analysis of Wheat GBSS1 Promoter:Tissue Specificity and DNA Methylation

The cis-regulatory elements of promoters regulate temporal and spatial expression of genes. DNA inethylation, histone methylation and histone acetylation are the main types of epigenetic modifications, which play important roles in plant growth and development. DNA methylation could seilenco transposons, affect gene imprinting and gene expression. In this study, we found that granule bound starch synthase 1 （GBSSI） gene is expressed specifically in wheat endosperm rath- er than in the embryo. We also analyzed the cis-elements within this promoter region and found some seed-specific elements. In order to confirm the tissue specifici- ty, we cloned 4k bp sequences upstream of GBSS1 gene to link to vector with GUS and this construct was transferred to tobacco by Agrobacterium mediated transfor- marion. The results showed that wheat GBSS1 promoter mediated the seed-specific expression of GUS gene, hut not mediated expression in embryo. In addition, we found that GBSSI promoter is methylated in wheat embryo and de-methylated in wheat endosperm. Our study might provide the molecular basis for specific expres- sion of GBSSI gene.