122 UNIQUE EXPRESSION PATTERNS OF DIFFERENTIATION, GROWTH, AND CELL STRUCTURE FACTORS IN THE ELONGATING PORCINE CONCEPTUS

2006 ◽  
Vol 18 (2) ◽  
pp. 170 ◽  
Author(s):  
L. A. Blomberg ◽  
J. R. Miles ◽  
K. A. Zuelke

Elongation of the trophectoderm and gastrulation of the embryonic disc, observed during gestational Days 11 (D11) through 12 (D12), denote a critical period of porcine conceptus development. Serial analysis of gene expression identified genes involved in cellular differentiation/structure (cytokeratin-8 and -18) and growth/cell migration/mesoderm-epithelial interaction (stratifin and midkine), which could potentially be regulated by steroids such as estrogen. Characterization of these factors is lacking in porcine conceptuses, therefore, the current study investigated mRNA expression of these factors in elongating conceptuses and primordial tissues as well as protein expression and cellular localization to better define their biological significance. Conceptuses examined were of ovoid (D11; 6-10 mm), tubular (D11; 11-50 mm), or filamentous (D12; >100 mm) morphology. Cells of the conceptus were highly proliferative at all stages and the embryonic disc of the ovoid conceptus was already polarized as indicated by the protein expression of Ki67 and brachyury. Real-time PCR was utilized to determine the transcript expression profiles. Differential expression of cytokeratin-18 and midkine were not apparent; however, cytokeratin-8 was clearly down-regulated in filamentous compared to ovoid conceptuses. In contrast, stratifin mRNA levels were greatest in tubular conceptuses of 42-50 mm size. Transcripts for cytokeratin-8 and -18, stratifin, and midkine were detected in both cell types (endoderm and trophoblast) of the trophoectoderm. Western blotting and/or immunohistochemistry were utilized to examine protein expression and cellular localization. The embryonic disc of ovoid conceptuses was almost devoid of cytokeratin-18 protein, however, its distribution was uniform throughout the trophectoderm at all stages of elongation. Stratifin and midkine proteins demonstrated more unique expression patterns within the conceptus. Distinct cell populations of the embryonic disc and the trophoectoderm contained stratifin; cellular localization was predominantly cytoplasmic but occasional nuclear translocation was evident. Furthermore, total protein levels of stratifin were not different among ovoid, tubular, and filamentous conceptuses, but proteolysis of the protein was apparent at the filamentous stage. Midkine protein expression was prominent in the embryonic disc of ovoid conceptuses. In tubular conceptuses, midkine was associated with cells that appeared to be migrating away from embryonic disc as well as some concentrating in the tips of the trophectoderm. Our findings suggested that cytokeratin-8 and -18 are associated primarily with the trophectoderm, as seen in other species. Furthermore, the distribution and localization of stratifin and midkine proteins could reflect attributed functions of these factors, minimal anti-proliferative activity in the rapidly growing conceptuses and cell migration important for gastrulation/trophectoderm elongation, respectively.

2009 ◽  
Vol 02 (02) ◽  
pp. 197-211 ◽  
Author(s):  
HONGLEI LI ◽  
XIAOGUANG CHEN ◽  
FUCHUN ZHANG ◽  
JI MA ◽  
CUNSHUAN XU

Cell migration plays the essential role in embryogenesis, wound healing, immunization, infection, and cancer metastasis. To study actions of cell migration-associated genes in liver regeneration, these genes were obtained by seaching for the related databases and literatures, and comprehensive analysis for the gene expression change during the regenerating process in liver was detected by Rat Genome 230 2.0 array, and identification of the LR-associated genes was through comparing the discrepancy in gene expression between sham operation and partial hepatectomy groups. The initially and totally expressed numbers of these genes in the initial phase of LR, G0/G1 transition, cell proliferation, cell differentiation, and structural-functional reconstruction were 88, 16, 79, 9, and 177, 90, 632, 207, respectively, illustrating that the associated genes triggered their expression mainly in the priming stage, and worked in different phases. Their expression similarity was classified into five groups and their expression patterns were categorized into 38 types, and the overall times of up-regulation and down-regulation were 589 and 427. Based on expression profiles and expression patterns of cell migration-associated genes in LR, it was confirmed that cell migration-associated genes rise in mRNA levels.


2003 ◽  
Vol 12 (2) ◽  
pp. 139-146 ◽  
Author(s):  
Mingzhang Guo ◽  
Mack H. Wu ◽  
Ferenc Korompai ◽  
Sarah Y. Yuan

The protein kinase C (PKC) pathway has recently been recognized as an important mechanism in the development of diabetic complications including cardiomyopathy and angiopathy. Although an increase in PKC kinase activity has been detected in the cardiovascular system of diabetic patients and animals, it is unclear whether the same pathological condition alters PKC at the transcriptional and translational levels. In this study we assessed quantitatively the mRNA and protein expression profiles of PKC isozymes in the heart and vascular tissues from streptozotocin-induced diabetic pigs. Partial regions of the porcine PKCα, β1, and β2 mRNAs were sequenced, and real-time RT-PCR assays were developed for PKC mRNA quantification. The results showed a significant increase in the mRNA levels of PKCα, β1, and β2 in the heart at 4–8 wk of diabetes. In concomitance, the PKCβ1 and β2 genes, but not the PKCα gene, were upregulated in the diabetic aorta. Correspondingly, there was a significant increase in the protein expression of PKCα and β2 in the heart and PKCβ2 in the aorta with a time course correlated to that of mRNA expression. In summary, PKCβ2 was significantly upregulated in the heart and aorta at both the transcriptional and translational levels during early stages of experimental diabetes, suggesting that PKCβ2 may be a prominent target of diabetic injury in the cardiovascular system.


2004 ◽  
Vol 16 (8) ◽  
pp. 763 ◽  
Author(s):  
Han-Seung Kang ◽  
Chae-Kwan Lee ◽  
Ju-Ran Kim ◽  
Seong-Jin Yu ◽  
Sung-Goo Kang ◽  
...  

In the present study, differential gene expression in the uteri of ovariectomised (OVX) and pro-oestrous rats (OVX v. pro-oestrus pair) was investigated using cDNA expression array analysis. Differential uterine gene expression in OVX rats and progesterone (P4)-injected OVX rats (OVX v. OVX + P4 pair) was also examined. The uterine gene expression profiles of these two sets of animals were also compared for the effects of P4 treatment. RNA samples were extracted from uterine tissues and reverse transcribed in the presence of [α32P]-dATP. Membrane sets of rat arrays were hybridised with cDNA probe sets. Northern blot analysis was used to validate the relative gene expression patterns obtained from the cDNA array. Of the 1176 cDNAs examined, 23 genes showed significant (>two-fold) changes in expression in the OVX v. pro-oestrus pair. Twenty of these genes were upregulated during pro-oestrus compared with their expression in the OVX rat uterus. In the OVX v. OVX + P4 pair, 22 genes showed significant (>two-fold) changes in gene expression. Twenty of these genes were upregulated in the OVX + P4 animals. The genes for nuclear factor I–XI, afadin, neuroligin 2, semaphorin Z, calpain 4, cyclase-associated protein homologue, thymosin β-4X and p8 were significantly upregulated in the uteri of the pro-oestrus and OVX + P4 rats of both experimental pairs compared with the OVX rat uteri. These genes appear to be under the control of P4. One of the most interesting findings of the present study is the unexpected and marked expression of the neuroligin 2 gene in the rat uterus. This gene is expressed at high levels in the central nervous system and acts as a nerve cell adhesion factor. According to Northern blot analysis, neuroligin 2 gene expression was higher during the pro-oestrus and metoestrus stages than during the oestrus and dioestrus stages of the oestrous cycle. In addition, neuroligin 2 mRNA levels were increased by both 17β-oestradiol (E2) and P4, although P4 administration upregulated gene expression to a greater extent than injection of E2. These results indicate that neuroligin 2 gene expression in the rat uterus is under the control of both E2 and P4, which are secreted periodically during the oestrous cycle.


2021 ◽  
Author(s):  
Leiyu Hao ◽  
Fengru Huang ◽  
Xinqian Yu ◽  
Bujie Xu ◽  
Yan Liu ◽  
...  

Abstract Background: Early growth response family members (EGRs), EGR1-4, have increasingly attracted attention in multiple cancers. However, the exact expression patterns and prognostic values of EGRs in the progress of breast cancer (BRCA) remain largely unknown. Methods: The mRNA expression and prognostic characteristics of EGRs were examined by the Cancer Genome Atlas (TCGA), Oncomine and Kaplan-Meier plotter. Enrichment analyses were conducted based on protein-protein interaction (PPI) network. The Tumor Immune Estimation Resource (TIMER) database and MethSurv were further explored. The protein expression level of EGR1 and cell migration were measured by Western blotting, immunohistochemistry, wound-healing assay and Boyden chamber assay in BRCA. Results: The transcriptional levels of EGR1/2/3 displayed significantly low expression in BRCA compared to that in normal tissues, while EGR4 was shown adverse expression pattern. Survival analysis revealed up-regulated EGR1-4 were remarkably associated with favorable relapse-free survival (RFS). A close correlation with specific tumor-infiltrating immune cells (TIICs) and several CpG sites of EGRs were exhibited. Immunohistochemistry assays showed that the protein expression of EGR1 was remarkably downregulated in BRCA compared to that in paracancerous tissues. Cell migration of MCF10A cells was increased after the silence of EGR1 by siRNA transfection.Conclusions: This study provides a novel insight to the role of EGR1 in the prognostic value and cell migration of BRCA.


BMC Genomics ◽  
2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Kejun Wang ◽  
Kaijie Yang ◽  
Qiao Xu ◽  
Yufang Liu ◽  
Wenting Li ◽  
...  

Abstract Background Embryonic mortality is a major concern in the commercial swine industry and primarily occurs early in gestation, but also during mid-gestation (~ days 50–70). Previous reports demonstrated that the embryonic loss rate was significant lower in Meishan than in commercial breeds (including Duroc). Most studies have focused on embryonic mortality in early gestation, but little is known about embryonic loss during mid-gestation. Results In this study, protein expression patterns in endometrial tissue from Meishan and Duroc sows were examined during mid-gestation. A total of 2170 proteins were identified in both breeds. After statistical analysis, 70 and 114 differentially expressed proteins (DEPs) were identified in Meishan and Duroc sows, respectively. Between Meishan and Duroc sows, 114 DEPs were detected at day 49, and 98 DEPs were detected at day 72. Functional enrichment analysis revealed differences in protein expression patterns in the two breeds. Around half of DEPs were more highly expressed in Duroc at day 49 (DUD49), relative to DUD72 and Meishan at day 49 (MSD49). Many DEPs appear to be involved in metabolic process such as arginine metabolism. Our results suggest that the differences in expression affect uterine capacity, endometrial matrix remodeling, and maternal-embryo cross-talk, and may be major factors influencing the differences in embryonic loss between Meishan and Duroc sows during mid-gestation. Conclusions Our data showed differential protein expression pattern in endometrium between Meishan and Duroc sows and provides insight into the development process of endometrium. These findings could help us further uncover the molecular mechanism involved in prolificacy.


Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2355-2355
Author(s):  
Steven M. Kornblau ◽  
David McCue ◽  
Kang L. Lu ◽  
Wenjing Chen ◽  
Kevin R. Coombes

Abstract Protein expression and activation determines the pathophysiology of leukemic cells in Myelodysplasia (MDS) and Acute Myelogenous Leukemia (AML) and is dependent on endogenous changes (e.g mutation, methylation) and exogenous signals from stromal interactions, cytokines (CTKN) and chemokines. We have previously performed proteomics on primary AML sample (using reverse phase protein arrays) and wanted to assess how cytokines affect protein expression and phosphorylation. Prior studies of CTKN expression in AML and MDS have generally measured individual CTKNs, but not provided an overall assessment of CTKN expression. We measured the level of 26 CTKN (IL-1RA, 1B, 2, 4 5, 6, 7 , 8 , 9, 10,12, 13, 15, 17, Eotaxin, FGFB, G-CSF, GM-CSF, IFNγ, IP10, MCP1, MIP1α, MIP1β, PDGF, TNFα and VEGF) using multiplex cytometry (Bioplex™, Biorad) in serum samples from 176 AML (138 untreated (New), 38 relapsed (REL)) and 114 MDS patients (97 New, 10 post biological therapy, 7 REL) and 19 normal (NL) subjects. Individual CTKN expression was not correlated with clinical features (e.g. age, gender, cytogenetics, FAB, HB, WBC, platelet etc). The levels of IL -1β, 4, 5, 6, 7,10,12, 13, 17, IFNγ, FGFB and MIP1α were significantly lower and IL-8 and 15 higher in AML/MDS compared to NL. The expression profiles of AML and MDS were statistically indistinguishable whether analyzed individually or by unsupervised hierarchical clustering, except for IL-8 and 13 (higher in AML) and VEGF (higher in MDS). When CTKN were evaluated individually in new AML cases higher levels of IL4, 5 and 10 correlated significantly with remission attainment, and higher levels of IL8, Il1Ra, IP-10, Mip1β, VEGF and ILR, correlated significantly with shorter survival. No CTKN predicted remission attainment or survival in MDS. Unsupervised hierarchical bootstrap clustering using Pearson correlation and average linkage of CTKN expression relative to other CTKN expression, where high levels of one CTKN correlated with high expression of the other, revealed 6 highly reproducible expression patterns: IL-1β 4, 7, 10, 12, 13, G-CSF, IFNγ, MIP1α and PDGF IL 1ra, 6, 8 Eotaxin, IP-10, MIP1β and VEGF, IL2, 9, 15 and GMCSF, IL5 IL-7, FGF-Basic, TNFα and MCP1. Similar unsupervised clustering of the samples based on CTKN expression using average linkage also revealed 5 disease clusters and a NL sample cluster (containing all 19 NL samples). Average expression levels of each CTKN in these 5 clusters show diminished expression of most CTKN that had high expression in the NL samples, with each group showing increase in expression in a distinct subset of CTKN relative to NL. Remission attainment was strongly associated with cytokine signature (P=0.005). Additional CTKN are being studied (SCF, TGFβ, IL3). Comparison of CTKN expression patterns with proteomic profiling of expression and phosphorylation status is pending. In summary, this is the largest sample set studied for multiple CTKN expression in AML and MDS and the first assessment of many of these CTKN in these diseases. Most CTKNs showed different expression in AML and MDS compared to NL. Interestingly, CTKN expression in AML and MDS were similar. Many CTKN are predictive of outcome individually. CTKN signatures distinguish groups of patients and are predictive of outcome. Correlation with proteomic profiling may suggest CTKN to target in combination with other targeted therapies to maximally affect activated pathways.


2006 ◽  
Vol 74 (7) ◽  
pp. 3853-3863 ◽  
Author(s):  
Sanghamitra Mukhopadhyay ◽  
Richard D. Miller ◽  
Erin D. Sullivan ◽  
Christina Theodoropoulos ◽  
Sarah A. Mathews ◽  
...  

ABSTRACT Chlamydia pneumoniae is an obligate intracellular pathogen that causes both acute and chronic human disease. Several in vitro models of chlamydial persistence have been established to mimic chlamydial persistence in vivo. We determined the expression patterns of 52 C. pneumoniae proteins, representing nine functional subgroups, from the gamma interferon (IFN-γ) treatment (primarily tryptophan limitation) and iron limitation (IL) models of persistence compared to those following heat shock (HS) at 42°C. Protein expression patterns of C. pneumoniae persistence indicates a strong stress component, as evidenced by the upregulation of proteins involved in protein folding, assembly, and modification. However, it is clearly more than just a stress response. In IFN persistence, but not IL or HS, amino acid and/or nucleotide biosynthesis proteins were found to be significantly upregulated. In contrast, proteins involved in the biosynthesis of cofactors, cellular processes, energy metabolism, transcription, and translation showed an increased in expression in only the IL model of persistence. These data represent the most extensive protein expression study of C. pneumoniae comparing the chlamydial heat shock stress response to two models of persistence and identifying the common and unique protein level responses during persistence.


Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3458-3458
Author(s):  
Tsz-Kwong Man ◽  
Mohammad Javad Najaf Panah ◽  
Jessica L. Elswood ◽  
Pavel Sumazin ◽  
Michele S. Redell

Abstract Introduction - Acute myeloid leukemia (AML) is an aggressive disease with a relapse rate of approximately 40% in children. Progress in improving cure rates has been slow, in part because AML is very heterogeneous. Molecular studies consistently show that most cases are comprised of distinct subclones that diminish or expand over the course of therapy. Single-cell profiling methods now allow parsing of the leukemic population into subsets based on gene and/or protein expression patterns. We hypothesized that comparing the features of the subsets that are dominant at relapse with those that are dominant at diagnosis would reveal mechanisms of treatment failure. Methods - We profiled diagnosis-relapse pairs from 6 pediatric AML patients by Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-Seq). All patients were treated at Texas Children's Cancer Center and consented to banking of tissue for research. CITE-Seq was performed by Immunai (New York, NY) using a customized panel of 65 oligonucleotide-tagged antibodies, the 10x Genomics Chromium system for single-cell RNA library generation, and the Novaseq 6000 for sequencing. After data cleanup and normalization, clustering by scRNA-seq was done using the Seurat package. Cell-type identification of clusters was facilitated by published healthy bone marrow scRNA-seq datasets (van Galen et al, Cell 2019). Differentially expressed genes (DEGs) and proteins (DEPs) between diagnosis and relapse were determined using Wilcoxin ranked sum tests. Results - We generated single-cell transcriptomes for a total of 28,486 cells from 12 samples, with a mean of 2373 cells and 1416 genes per sample. Samples were integrated with batch effect correction, producing 30 distinct clusters (cell types) in total (Figure 1A). Cell types with expression profiles consistent with lymphocytes and erythroid precursors were identified in multiple patients, whereas AML cell types tended to be specific to individual patients (Figure 1B). For patients TCH1, TCH2 and TCH3, the most abundant cell types at diagnosis were rare at relapse, and cell types that were rare at diagnosis became dominant at relapse. For these 3 cases, we identified DEGs between the dominant diagnosis cell types and dominant relapse cell types. We found 18 genes that were upregulated at relapse in at least 2 of the cases. Several genes related to actin polymerization were enriched (ARPC1B, ACTB, PFN1), possibly reflecting an enhanced capacity for adhesion and migration. Also of note, macrophage migration inhibitory factor (MIF) and its receptor CD74 were upregulated at relapse, suggesting a role in chemoresistance. For patients TCH4, TCH5 and TCH6, the same cell types that were abundant at diagnosis were also abundant at relapse, and few genes were significantly altered between diagnosis and relapse in multiple cases. Only SRGN, which encodes the proteoglycan serglycin, and GAPDH were altered in 2 of these 3 cases, and both were downregulated at relapse. We performed similar comparisons to identify proteins that were differentially expressed between diagnosis and relapse pairs. The number of DEPs between the dominant diagnosis and relapse cell types ranged from 0 (TCH1 and TCH6) to 5 (TCH2). The only protein altered in more than one case was CD7, which was enriched at relapse in TCH2, TCH3 and TCH4. Conclusions - From CITE-Seq profiling of 6 pediatric AML cases we identified two distinct patterns of relapse. For 3 cases, relapse occurred by expansion of a subset that was small but present at diagnosis. Enrichment of genes associated with adhesion and survival signaling suggests that these cells survived because they were well-equipped to take advantage of interactions with the microenvironment. For 3 other cases, the population that was dominant at diagnosis persisted and expanded at relapse with few substantial changes in gene or protein expression profiles. This pattern suggests that these AML cells were a priori equipped to survive chemotherapy, even though bulk disease levels were transiently reduced below the limit of detection. Most profiled proteins did not change substantially between diagnosis and relapse. An exception is CD7, which was enriched at relapse in 50% of our cases and represents a potential therapeutic target. Analysis of more cases will refine these relapse patterns, reveal potential mechanisms of chemoresistance and inform the development of novel therapies. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.


2016 ◽  
Author(s):  
Daniel A. Pollard ◽  
Ciara K. Asamoto ◽  
Homa Rahnamoun ◽  
Austin S. Abendroth ◽  
Suzanne R. Lee ◽  
...  

ABSTRACTHeritable variation in gene expression patterns plays a fundamental role in trait variation and evolution, making understanding the mechanisms by which genetic variation acts on gene expression patterns a major goal for biology. Both theoretical and empirical work have largely focused on variation in steady-state mRNA levels and mRNA synthesis rates, particularly of protein-coding genes. Yet in order for this variation to affect higher order traits it must lead to differences at the protein level. Variation in protein-specific processes including protein synthesis rates and protein decay rates could amplify, mask, or even reverse effects transmitted from the transcript level, but the extent to which this happens is unclear. Moreover, mechanisms that underlie protein expression variation under dynamic conditions have not been examined. To address this challenge, we analyzed how mRNA and protein expression dynamics covary between two strains ofSaccharomyces cerevisiaeduring mating pheromone response. Although divergentsteady-statemRNA expression levels explained divergentsteady-stateprotein levels for four out of five genes in our study, the same was true for only one out of five genes for expressiondynamics. By integrating decay rate and allele-specific protein expression analyses, we resolved that expression divergence for Fig1p was caused by genetic variation acting intranson protein synthesis rate, expression divergence for Ina1p was caused bycis-by-transepistatic effects on transcript level and protein synthesis rate, and expression divergence for Fus3p and Tos6p were caused by divergence in protein synthesis rates. Our study demonstrates that steady-state analysis of gene expression is insufficient to understand the impact of genetic variation on gene expression variation. An integrated and dynamic approach to gene expression analysis - comparing mRNA levels, protein levels, protein decay rates, and allele-specific protein expression - allows for a detailed analysis of the genetic mechanisms underlying protein expression divergences.


Endocrinology ◽  
2011 ◽  
Vol 152 (3) ◽  
pp. 989-1000 ◽  
Author(s):  
Monika Proszkowiec-Weglarz ◽  
Stacy E. Higgins ◽  
Tom E. Porter

The anterior pituitary gland plays an important role in the regulation of many physiological processes. Formation of Rathke's pouch (RP), the precursor of the anterior pituitary, involves evagination of the oral ectoderm in a multi-step process regulated by cell interactions, signaling pathways, and transcription factors. Chickens are an excellent model to study development because of the availability of large sample sizes, accurate timing of development, and embryo accessibility. The aim of this study was to quantify mRNA expression patterns in the developing chicken anterior pituitary to evaluate the chicken embryo as a model for mammalian pituitary development. The expression profiles of 16 genes differentially expressed in RP and neuroectoderm were determined in this study. Among these, Pitx1, Pitx2, and Hesx1 mRNA levels were high on embryonic days (e) 2.5 to e3 in RP and decreased during development. Expression of Pit1 and Tbx19 mRNA in RP reached the highest levels by e7 and e6.5, respectively. Levels of glycoprotein subunit α mRNA increased beginning at e4. FGF8 mRNA showed the highest expression at e3 to e3.5 in neuroectoderm. BMP2 showed slight decreases in mRNA expression in both tissues during development, while Isl1 and Noggin mRNA expression increased in later development. Taken together, we present the first quantitative transcriptional profile of pituitary organogenesis. Our results will help further understanding of the functional development of this gland. Moreover, because of the high similarity in gene expression patterns observed between chicken and mouse, chickens could serve as an excellent model to study genetic and molecular mechanisms underlying pituitary development.


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