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Quantitative evaluation of tissue-specific cell adhesion at the level of a single cell pair.

Tissue specificity of cell adhesion was directly characterized in a unit cell interaction using a novel laser trapping cell manipulator in combination with a fixed micropipet. We quantified the adhesive specificity of endodermal and ectodermal epithelial cells from Hydra, which are known to sort out within hours after being dissociated and then randomly reaggregate. It was shown that homotypic pairs of cells from the same tissue source could adhere to each other within a certain period, while heterotypic pairs could not form an adhesion. It was also found that the adhesion probability was higher in endodermal epithelial cell pairs than in ectodermal epithelial cell pairs. The former pairs could adhere with a contact period of less than 30 sec, while 60% of the latter remained nonadherent even after a 6-min forced contact. The adhesive strength of the latter was estimated to be as large as 30 pN, while that of the former was much larger than 50 pN. The tissue-specific adhesivity quantitatively measured provides a new insight into the mechanism of cell sorting.

Animals↗

Tissue-specific characteristics of cytotoxic graft-infiltrating T cells during renal allograft rejection.

BACKGROUND: The alloresponse after renal transplantation was studied using alloreactive T cells generated in vivo (from renal biopsies) and in vitro (from mixed kidney lymphocyte cultures). METHODS: Tissue specificity of graft-infiltrating T cells (GIC) was investigated using donor-derived proximal tubular epithelial cells (PTEC) and splenocytes as targets in cytotoxicity assays. RESULTS: The outgrowth of cytotoxic T cells was associated with histologically proven interstitial rejection. GIC were categorized into four groups: (1) GIC cytotoxic for both PTEC and splenocytes (n=30), (2) noncytotoxic GIC (n=8), (3) GIC recognizing only splenocytes (n=1), and (4) GIC specifically recognizing PTEC (n=7). Similar tissue-specific T cells could be generated in vitro using mixed kidney lymphocyte cultures. Cytotoxicity of GIC from biopsies with moderate to severe rejection was CD8 independent, whereas cytotoxicity toward splenocytes was CD8 dependent. CONCLUSIONS: Our results show that polyclonal cytotoxic T-cell responses with tissue-specific characteristics are elicited during rejection episodes after renal transplantation.

CD3 Complex↗

[Tissue-specific placental mosaicism for autosomal trisomies in spontaneous human abortuses: mechanisms of formation and phenotypic effects].

The frequencies of autosomal trisomies in extraembryonic human tissues were estimated in the cases of different abnormalities of prenatal development, from the confined placental mosaicism (CPM) with either relatively normal embryogenesis or restricted intrauterine growth to spontaneous abortion. A tissue-specific compartmentalization was found to be characteristic of cell lines with trisomies for individual autosomes. Analysis of various phenotypical effects of chromosomal aberrations associated with mosaicism is necessarily required to understand the mechanisms and factors responsible for tissue chromosomal mosaicism. Based on analysis of the cell karyotype during prenatal diagnosing of chromosome aberrations in tissues of both extraembryonic and embryonic origin, in 1996, Wolstenholme proposed a model of CPM for individual chromosomes. According to the model, the distribution of cell lines with autosomal trisomies between extraembryonic tissues depends on the ratio between meiotic and mitotic mutations early in embryonic development. However, the model cannot be used to study tissue chromosomal mosaicism in spontaneous abortions, because little information is available on cell karyotype in embryonic tissues themselves after intrauterine fetal death. In this work, a model of tissue-specific chromosomal mosaicism was suggested based on the data on cell karyotype determined in extraembryonic tissues alone, which can be helpful in evaluating the contribution of tissue chromosomal differences into the etiology of early intrauterine death. Along with the experimental evidence, comparative analysis of the two models indicated that the meiotic chromosome nondisjunction plays the major role in trisomy formation and the resultant spontaneous arrest of embryonic development. Other factors responsible for tissue-specific distribution of chromosomal aberrations are also discussed. These are differences in cell proliferative activity, as well as changes in compartmentalization and migration of cells with abnormal karyotypes.

Abortion, Spontaneous↗

DNA replication is required for tissue-specific enzyme development in ascidian embryos.

Tyrosinase which is a tissue-specific enzyme in the pigment cells of the brain of the ascidian embryo, is thought to be synthesized with activation of appropriate genes, and the enzyme synthesis begins at the early tailbud stage. If embryos at early cleavage stages up to the 64-cell stage are continuously treated with aphidicolin (a specific inhibitor of DNA synthesis), cleavage of the embryos is arrested and they do not differentiate the enzyme. However, the early gastrulae and embryos at later stage that have been permanently arrested with aphidicolin do produce the enzyme. Alkaline phosphatase, a tissue-specific enzyme of the endodermal cells, has been shown to be synthesized by a preformed maternal mRNA and is first detected histochemically at the late gastrula stage. If embryos at early cleavage stages up to the 16-cell stage are prevented from undergoing further divisions with aphidicolin, the arrested embryos do not form the enzyme. However, embryos at the 32-cell and later stages that have been permanently arrested with aphidicolin are able to differentiate the enzyme activity. These results suggest that several DNA replications are required for the histospecific enzyme development in ascidian embryos.

Alkaline Phosphatase↗

Competition of PTB with TIA proteins for binding to a U-rich cis-element determines tissue-specific splicing of the myosin phosphatase targeting subunit 1.

A considerable amount of smooth muscle phenotypic diversity is generated by tissue-specific and developmentally regulated splicing of alternative exons. The control mechanisms are unknown. We are using a myosin phosphatase targeting subunit-1 (MYPT1) alternative exon as a model to investigate this question. In the present study, we show that the RNA binding proteins TIA and PTB function as antagonistic enhancers and suppressors of splicing of the alternative exon, respectively. Each functions through a single U-rich element, containing two UCUU motifs, just downstream of the alternative exon 5' splice site. Tissue-specific down-regulation of TIA protein in the perinatal period allows PTB to bind to the U-rich element and suppress splicing of the alternative exon as the visceral smooth muscle acquires the fast-phasic smooth muscle contractile phenotype. This provides a novel role for PTB in the tissue-specific regulation of splicing of alternative exons during the generation of smooth muscle phenotypic diversity.

Alternative Splicing↗

Cytolytic T lymphocytes from human renal allograft biopsies are tissue specific.

The cytolytic activity of T lymphocytes infiltrating renal allografts from recipients undergoing episodes of acute cellular rejection was studied. These T-cell populations, composed of both CD4+ and CD8+ cells, demonstrated significant cytolytic activity against both donor-derived KCLs and B-LCLs. In five of 21 biopsy-derived lines greater cytolytic activity was measured against donor KCLs compared to donor B-LCLs, suggesting the presence of kidney antigen-specific, MHC-restricted clones. Clones developed by stimulation with donor B-LCLs lysed both donor B-LCLs and KCLs while clones developed on donor KCLs as stimulator cells showed tissue specificity. Three of 26 clones recognized tissue-specific antigens in the context of donor MHC class I antigens lysing donor KCLs but not B-LCLs. These data demonstrate that a subpopulation of T cells recognizing kidney-specific antigens are present in biopsies of renal allograft recipients undergoing acute cellular rejection. This subpopulation of tissue-specific cytotoxic T lymphocytes may prove to contribute significantly to the pathology of allograft rejection.

Biopsy↗

Gene-Ontology analysis reveals association of tissue-specific 5' CpG-island genes with development and embryogenesis.

A key open question in the understanding of the biology of DNA methylation relates to the origin and function of CpG islands, stretches of GC-rich and relatively CpG-rich DNA sequence that often colocalize with promoter regions. All housekeeping, but also a substantial minority of tissue-specific genes are associated with the CpG islands. Limited experimental evidence suggests that CpG islands are associated with promoters or replication origins active during early development. Although this hypothesis is attractive for widely expressed genes, which would be expected to be expressed during early development, many tissue-specific genes also contain CpG islands. In this work, we used a genome-wide Gene-Ontology (GO)-based approach to analyze associations between GO terms and the presence of 5' CpG islands in human Reference Sequence (RefSeq) genes. We found that 19 of the 3849 GO terms with at least one annotated human sequence showed a highly significant association with the likelihood of 5' CpG islands being present in genes annotated to that term. In particular, the term 'development' showed a highly significantly increased proportion of 5' CpG island genes. The overrepresentation of 5' CpG island genes was even more significant for tissue-specific RefSeqs annotated to development as well as many of its descendent terms. In addition, the proportion of expressed sequence tags from embryonic libraries amongst tissue-specific genes was twice as high for RefSeqs with 5' CpG islands as for those without CpG islands. These results provide strong support for previous speculations that early embryonic expression is associated with CpG islands.

Animals↗

Mechanisms in tissue-specific regulation of estrogen biosynthesis in humans.

In humans, aromatase P450, which catalyses conversion of C(19)-steroids to estrogens, is expressed in several tissues, including gonads, brain, adipose tissue, skin and placenta, and is encoded by a single-copy gene (CYP19); however, this does not hold true for all species. The human gene is approximately 130 kb and its expression is regulated, in part, by tissue-specific promoters and by alternative splicing mechanisms. Using transgenic mouse technology, it was observed that ovary-, adipose tissue- and placenta-specific expression of human CYP19 is directed by relatively small segments of DNA within 500 bp upstream of each of the tissue-specific first exons. Thus, the use of alternative promoters allows greater versatility in tissue-specific regulation of CYP19 expression. Characterization and identification of transcription factors and crucial cis-acting elements within genomic regions that direct tissue-specific expression will contribute to improved understanding of the regulation of CYP19 expression in the tissues that synthesize estrogens under both physiological and pathophysiological conditions.

Adipose Tissue↗

Meta-analysis discovery of tissue-specific DNA sequence motifs from mammalian gene expression data.

BACKGROUND: A key step in the regulation of gene expression is the sequence-specific binding of transcription factors (TFs) to their DNA recognition sites. However, elucidating TF binding site (TFBS) motifs in higher eukaryotes has been challenging, even when employing cross-species sequence conservation. We hypothesized that for human and mouse, many orthologous genes expressed in a similarly tissue-specific manner in both human and mouse gene expression data, are likely to be co-regulated by orthologous TFs that bind to DNA sequence motifs present within noncoding sequence conserved between these genomes. RESULTS: We performed automated motif searching and merging across four different motif finding algorithms, followed by filtering of the resulting motifs for those that contain blocks of information content. Applying this motif finding strategy to conserved noncoding regions surrounding co-expressed tissue-specific human genes allowed us to discover both previously known, and many novel candidate, regulatory DNA motifs in all 18 tissue-specific expression clusters that we examined. For previously known TFBS motifs, we observed that if a TF was expressed in the specified tissue of interest, then in most cases we identified a motif that matched its TRANSFAC motif; conversely, of all those discovered motifs that matched TRANSFAC motifs, most of the corresponding TF transcripts were expressed in the tissue(s) corresponding to the expression cluster for which the motif was found. CONCLUSION: Our results indicate that the integration of the results from multiple motif finding tools identifies and ranks highly more known and novel motifs than does the use of just one of these tools. In addition, we believe that our simultaneous enrichment strategies helped to identify likely human cis regulatory elements. A number of the discovered motifs may correspond to novel binding site motifs for as yet uncharacterized tissue-specific TFs. We expect this strategy to be useful for identifying motifs in other metazoan genomes.

Algorithms↗

Two-dimensional and three-dimensional time-lapse microscopic magnetic resonance imaging of Xenopus gastrulation movements using intrinsic tissue-specific contrast.

The amphibian embryo undergoes radical tissue transformations during blastula and gastrula stages, but live observation of internal morphogenetic events by optical microscopy is not feasible due to the opacity of the early embryo. Here, we report on the use of microscopic magnetic resonance imaging (MRI) to directly follow morphogenetic movements during blastula and gastrula stages of the Xenopus laevis embryo. We compare three different MRI modalities that take advantage of the intrinsic contrast present in embryonic tissues: three-dimensional (3D) fat-imaging, 3D water-imaging, and 2D high-speed high-resolution imaging of early embryonic stages. We show that the features revealed by the intrinsic contrast correlate with the histological structure of the embryo. Using this tissue specific intrinsic contrast, the main embryonic tissues and internal tissue movements as well as archenteron invagination can be differentiated without cell labeling. We present 2D and 3D time-lapse sequences of early Xenopus embryonic development, spanning the stages from early blastula to the end of gastrula, which show the complex internal rearrangements of gastrulation in essentially real-time.

Animals↗

Glucocorticoids control phosphoenolpyruvate carboxykinase gene expression in a tissue specific manner.

Cytosolic Phosphoenolpyruvate carboxykinase is a key gluconeogenic enzyme which is expressed in a tissue specific manner in the liver, kidney and adipose tissue and is under hormonal control. The effect of glucocorticoids on expression of the gene coding for phosphoenolpyruvate carboxykinase in adipose tissue has been studied in vivo in rats and in vitro in adipose tissue organ culture and mouse 3T3 L1 adipocytes. Glucocorticoids, both in vivo and in vitro, repress the steady state level of phosphoenolpyruvate carboxykinase mRNA in the adipose tissue while increasing it in the kidney. The size of the mRNA and its 5' end are identical in adipose tissue and kidney, thus the same promoter is used in all tissues. The inhibitory effect of glucocorticoids on phosphoenolpyruvate carboxykinase gene expression was located at the level of transcription. As glucocorticoids are known to stimulate transcription of phosphoenolpyruvate carboxykinase gene in the liver and kidney, the inhibitory effect on its transcription in adipose tissue suggests that tissue specific transcription factors may modulate the effect of glucocorticoids.

Adipose Tissue↗

Antioxidant systems of the avian embryo: tissue-specific accumulation and distribution of vitamin E in the turkey embryo during development.

Tissue-specific accumulation of tocopherols and tocotrienols in turkey tissues during embryonic development and their susceptibility to lipid peroxidation were investigated. Fertile turkey eggs were incubated using standard commercial conditions. Embryonic tissues were collected at 16, 22, 25 d of incubation and from day-old poults (referred to as day 29) and alpha-; beta- + gamma- and delta-tocopherols and respective tocotrienols were analysed by HPLC. A turkey diet provided to the parent hens contained the complete range of tocopherols and tocotrienols. Between days 16 and 22 of embryo development, the alpha-tocopherol concentration in the liver remained constant and then increased significantly (P<0.01) reaching a maximum just after hatching. Similar changes were observed for the other tocopherols and tocotrienols. The accumulation of alpha-tocopherol in the yolk sac membrane (YSM) started after day 20 of development and at hatching the alpha-tocopherol concentration in the YSM was twice that of beta- + gamma-tocopherols and 15 times greater than that of alpha-tocotrienol. In the kidney, heart, lung, muscle and adipose tissues a gradual increase in tocopherol and tocotrienol concentrations took place between days 20 and 25 of development with a sharp increase in particular of alpha-tocopherol between days 25 and 29. There was a discrimination between tocopherols and tocotrienols during their assimilation from the diet by the parent hen and during metabolism by the developing turkey embryo. Tissue-specific features in the susceptibility to lipid peroxidation were found with the brain being the most susceptible to lipid peroxidation at day 25 and in day-old poults.

Animal Feed↗

Population genetics of Drosophila amylase. I. Genetic control of tissue-specific expression in D. pseudoobscura.

Drosophila pseudoobscura is polymorphic for tissue-specific expression of alpha-amylase in adult midguts. This enzyme is encoded by a single locus, Amy, on the third chromosome. In this paper we show: (1) Up to about 12 days post-eclosion, the midgut activity patterns remain stable; after 12 days areas not showing activity previously begin to show activity. Thus, the genes controlling the expression of Amy are temporally acting. (2) Diet affects the quantitative, but not the qualitative, expression of Amy. (3) The expression of Amy in adult midguts is under genetic control. Selection for different frequencies of patterns is possible; realized heritabilities are 0.20 to 0.50. Partial linkage with third chromosome inversions has been demonstrated; the genes or elements controlling Amy expression are not, however, confined to the third chromosome. (4) The genetic elements controlling tissue-specific expression of amylase do not coordinately control the expression of five other "digestive-type" enzymes that were studied.--This polymorphism appears to be analogous to that studied by Abraham and Doane (1978) in D. melanogaster, wherein they have mapped regulatory genes.

Amylases↗

Determination of tissue specificity of the enhancer by combinatorial operation of tissue-enriched transcription factors. Both HNF-4 and C/EBP beta are required for liver-specific activity of the ornithine transcarbamylase enhancer.

The enhancer of the rat ornithine transcarbamylase gene is located 11 kilobases upstream from the transcription start site and has been shown to be hepatoma cell-specific. Using transgenic mice, we showed that this enhancer is capable of activating transcription in a liver-specific manner, inverting the tissue specificity of the homologous promoter that is by itself more active in the small intestine than in the liver. Transient transfection analysis with cultured hepatoma cells indicated that the enhancer activity resides in the approximately 110-base pair region containing four protein-binding sites, two for hepatocyte nuclear factor-4 (HNF-4) and two for CCAAT/enhancer binding protein (C/EBP), both of which are liver-selective transcription factors. Concatemerization of a region containing one HNF-4 and one C/EBP site led to reconstitution of the hepatoma cell-specific enhancer, and intactness of these two sites was strictly required for the enhancer activity. Furthermore, cotransfection experiments showed that both HNF-4 and C/EBP beta are necessary, and neither alone sufficient, for activation of the reconstituted enhancer in nonhepatic cells. Requirement of combinatorial operation of at least two liver-enriched transcription factors for transcriptional activation successfully explains why these liver-selective but not strictly liver-specific factors can confer more restricted liver specificity on transcription of their target genes.

Animals↗

Tissue-specific activity of the pro-opiomelanocortin gene promoter.

The pro-opiomelanocortin (POMC) gene is specifically expressed in corticotroph cells of the anterior pituitary. To define the POMC promoter sequences responsible for tissue-specific expression, we assessed POMC promoter activity by gene transfer into POMC-expressing pituitary tumor cells (AtT-20) and fibroblast L cells. The rat POMC promoter was only efficiently utilized and correctly transcribed in AtT-20 cells. 5'-End deletion analysis revealed two promoter regions required for activity in AtT-20 cells. When tested by fusion to a heterologous promoter, DNA fragments corresponding to both regions exhibited tissue-specific activity, suggesting the presence of at least two tissue-specific DNA sequence elements within the promoter. In summary, POMC promoter sequences from -480 to -34 base pairs appear sufficient to mimic the specificity of anterior pituitary expression.

Animals↗

Identification of tissue-specific DNase I hypersensitive sites in the rabbit flavin-containing monooxygenase form 2 gene.

Previous studies demonstrated that the flavin-containing monooxygenases (FMO) are expressed in a tissue-specific manner. To begin an elucidation of mechanisms regulating this expression pattern, genomic clones for rabbit FMO2 were isolated and characterized. Two clones were isolated from a lambda EMBL3 genomic library and shown to span approximately 30 kilobase pairs but to contain only about 800 base pairs of coding information. Primer extension analysis was used to map the transcription start site, extending the previously published cDNA sequence by 17 base pairs. The FMO2 promoter does not utilize a classical TATA box, nor are HTF islands present (DNA domains rich in cleavable sites for 5-methyl-cytidine/guanosine-sensitive restriction enzymes). Rather, homology with promoters controlled by initiator elements is observed. Previous studies demonstrated FMO2 expression in rabbit pulmonary Clara and type II cells [Overby, Nishio, Lawton, Plopper, and Philpot: Exp. Lung Res. 18, 131, (1992)]. In the present study, a highly enriched Clara/type II cell population was prepared, and the FMO2 gene was analyzed for DNase I-hypersensitive and methylated regions. These data were then contrasted with those obtained from a similar analysis of the nonexpressed, hepatocyte FMO2 gene. No difference in the methylation status was observed. However, Clara/type II cell-specific DNase I-hypersensitive sites are located within the promoter region of the FMO2 gene. Thus, tissue-specific transcription factors likely are more prominent than methylation in regulating FMO2 expression. Consistent with this observation, both polyomavirus enhancer activator 3 (E26 transformation specific) and thyroid transcription factor 1 consensus sequences are present within the tissue-specific DNase I-hypersensitive domain.

Animals↗

In vivo ultrasonographic exposimetry: human tissue-specific attenuation coefficients in the gynecologic examination.

OBJECTIVE: The purpose of the current study was to determine in vivo, tissue-specific ultrasonic attenuation coefficients for each of the tissue layers comprising the anterior abdominal wall, uterus, and vagina with use of a quantitative multilayer tissue model. We wanted to validate the "homogeneous" tissue model-based Food and Drug Administration derating factor of 0.3 dB/cm-MHz applied to obstetric-use ultrasonography systems. STUDY DESIGN: With use of a 3. 0-MHz mechanical sector scanner and our previously tested exposimetry equipment, we obtained a set of at least 5 separate acoustic pressure waveforms from each test subject by placing a calibrated 7-element linear-array hydrophone in the anterior vaginal fornix while she was undergoing transabdominal ultrasonography. Corresponding sets of reference in vitro acoustic pressure waveforms were also recorded for each test subject in a 37 degrees C water bath. All linear measurements of individual layer thicknesses and total distances were made on-line with use of electronic calipers. A set of multiple and independent insertion loss values, denoted ILn, was calculated for path n between the abdominal surface and the hydrophone from n sonograms for each test subject. Each tissue layer type was identified and its thickness along each path n was measured. The thickness of tissue type m along path n was denoted by dnm. The only unknown quantities left were the attenuation coefficients Am of each of the m tissue layers for that test subject. The overestimated set of equations dnm Am = ILn was solved for Am with use of a nonnegative least-squares solution technique. RESULTS: With use of data from 162 independent insertion loss estimate paths, the overall tissue-specific attenuation coefficients for each of the tissue layer types, expressed as mean value +/- SD, were 2.3 +/- 1.5 dB/cm-MHz for the skin and subcutaneous layer, 3.1 +/- 2.5 dB/cm-MHz for skeletal muscle, 0.6 +/- 0.5 dB/cm-MHz for myometrium, and 3.6 +/- 2.7 dB/cm-MHz for the vaginal wall. The overall insertion loss assuming the "homogeneous" tissue model was 0.7 +/- 0.3 dB/cm-MHz. CONCLUSIONS: We have determined the specific ultrasonic attenuation coefficients for each of the tissue layers comprising the anterior abdominal wall, uterus, and vagina and validated the Food and Drug Administration derating factor of 0.3 dB/cm-MHz applied to obstetric use ultrasonography systems. Of all the models proposed, the "homogeneous" tissue model appears to be the best model for determining ultrasonic exposure risk during reproductive ultrasonographic examinations.

Abdomen↗

A tandemly repeated thyroglobulin core promoter has potential to enhance efficacy for tissue-specific gene therapy for thyroid carcinomas.

Recombinant adenoviruses, carrying herpes simplex virus thymidine kinase (HSVtk) genes, were developed to evaluate the possibility of tissue-specific gene therapy for thyroid carcinomas. The HSVtk gene was driven by a minimal thyroglobulin (TG) promoter (AdTGtk) and a tandemly repeated minimal TG promoter (Ad2 x TGtk) to obtain thyroid-specific cell killing ability. The transduction of HSVtk genes by infection with Ad2 x TGtk followed by ganciclovir (GCV) treatment showed more powerful cytotoxicity for TG-producing FRTL5 cells, a rat normal thyroid cell line, and FTC-133 cells, a human follicular thyroid carcinoma cell line, than when infected with AdTGtk in vitro. The cell killing ability of Ad2 x TGtk was 10- to 30-fold higher than that of AdTGtk and similar to that of AdCMVtk, which carries HSVtk under the control of CMV promoter. Whereas after treatment with adenovirus/GCV to non-TG-producing cell lines (undifferentiated thyroid carcinoma cell lines and carcinoma cell lines from other tissues), Ad2 x TGtk and AdTGtk needed more than 100-fold concentrated GCV to reach IC(50) compared to AdCMVtk. We confirmed the enhanced efficacy of Ad2 x TGtk for tissue-specific cytotoxicity in vivo. After adenovirus/GCV treatment for FTC-133 tumor-bearing nude mice, Ad2 x TGtk enhanced tumor growth inhibition and survival rates compared to AdTGtk. Tumor growth inhibition and survival rates by Ad2 x TGtk were similar to that by AdCMVtk. Moreover, any toxic effect for rat normal tissues was not revealed after intravenous injections with Ad2 x TGtk and intraperitoneal administrations with GCV in vivo, whereas severe liver damages were observed after treatment with AdCMVtk/GCV. These data indicate a beneficial effect of Ad2 x TGtk for tissue-specific gene therapy for TG-producing thyroid carcinomas without toxicity for normal tissues.

Adenoviridae↗