scholarly journals Colonic Handling of Calcium Appears To Occur Via Different Pathways in Mouse Models of Infants and Adults

2021 ◽  
Vol 5 (Supplement_2) ◽  
pp. 1305-1305
Author(s):  
Megan Beggs ◽  
R Todd !Alexander
Keyword(s):  
Gene Expression ◽  
Molecular Mechanisms ◽  
Epithelial Transport ◽  
Engineering Research ◽  
Ussing Chambers ◽  
Funding Sources ◽  
Critical Age ◽  
Future Work

Abstract Objectives Calcium (Ca2+) is a vital micronutrient for many physiological functions with the greatest rate of accumulation occurring during the critical period of infancy. Previous work has demonstrated that molecular mechanisms of intestinal Ca2+ absorption across the small intestine are significantly different in animal models of infants and adults to permit greater absorption early in life. The colon contributes to overall Ca2+ balance in adults via transcellular, TRPV6 mediated and paracellular claudin-2 and -12 mediated pathways. Whether these same colonic pathways contribute to overall Ca2+ absorption in infants is not known. Here we aimed to investigate the molecular details of Ca2+ absorption across the large intestine in murine models of infancy relative to older mice. Methods Mice at 14 days (P14) were used as a model of suckling infants and mice at 2 months were employed to represent adult physiology. Wildtype and mutant mice with a non-functioning Trpv6 or deletions of Cldn2 or Cldn12 were used. Net 45Ca2+ flux (JCa) and Ca2+ permeability (PCa) were measured in Ussing chambers. Gene expression was determined by real-time PCR. Results JCa indicates net absorption across the colon at both P14 and 2 months. While gene expression of Trpv6 and S100g suggest greater cellular uptake of Ca2+ into colonocytes at P14, net JCa in vitro was not different than at 2 months. In contrast to previous work in mice at 2 months, TRPV6 does not mediate JCa at P14. PCa was 20% greater at P14 than 2 months, suggesting greater capacity for bidirectional diffusion of Ca2+ down an electrochemical gradient in younger mice. In contrast to previous work in mice at 2 months, claudin-2 and claudin-12 do not mediate PCa at P14 and, expression of Cldn2 and Cldn12 were significantly reduced in younger mice. Conclusions These results improve our understanding of intestinal Ca2+ handling during a critical age early in life. Future work is required to delineate molecular details under in vivo conditions of colonic Ca2+ transport in infants. Funding Sources This work was funded by grants from the Women and Children's Health Research Institute, which is supported by the Stollery Children's Hospital Foundation, and the National Sciences and Engineering Research Council to RTA, who is the Canada Research Chair in Renal Epithelial Transport Physiology.

Serial analysis of gene expression (SAGE) during porcine embryo development

2004 ◽  
Vol 16 (2) ◽  
pp. 87 ◽  
Author(s):  
Le Ann Blomberg ◽  
Kurt A. Zuelke
Keyword(s):  
Gene Expression ◽  
Functional Genomics ◽  
Embryo Development ◽  
Molecular Mechanisms ◽  
Physiological Role ◽  
Transgenic Animals ◽  
Specific Gene ◽  
Research Strategies

Functional genomics provides a powerful means for delving into the molecular mechanisms involved in pre-implantation development of porcine embryos. High rates of embryonic mortality (30%), following either natural mating or artificial insemination, emphasise the need to improve the efficiency of reproduction in the pig. The poor success rate of live offspring from in vitro-manipulated pig embryos also hampers efforts to generate transgenic animals for biotechnology applications. Previous analysis of differential gene expression has demonstrated stage-specific gene expression for in vivo-derived embryos and altered gene expression for in vitro-derived embryos. However, the methods used to date examine relatively few genes simultaneously and, thus, provide an incomplete glimpse of the physiological role of these genes during embryogenesis. The present review will focus on two aspects of applying functional genomics research strategies for analysing the expression of genes during elongation of pig embryos between gestational day (D) 11 and D12. First, we compare and contrast current methodologies that are being used for gene discovery and expression analysis during pig embryo development. Second, we establish a paradigm for applying serial analysis of gene expression as a functional genomics tool to obtain preliminary information essential for discovering the physiological mechanisms by which distinct embryonic phenotypes are derived.

scholarly journals Retinoic acid activates interferon regulatory factor-1 gene expression in myeloid cells

Blood ◽  
1996 ◽  
Vol 88 (1) ◽  
pp. 114-123 ◽  
Author(s):  
S Matikainen ◽  
T Ronni ◽  
M Hurme ◽  
R Pine ◽  
I Julkunen
Keyword(s):  
Gene Expression ◽  
Retinoic Acid ◽  
Growth Inhibition ◽  
Molecular Mechanisms ◽  
Gene Activation ◽  
Oligoadenylate Synthetase ◽  
Nb4 Cells ◽  
Drug Of Choice

All-trans-retinoic acid (ATRA) is the drug of choice in the treatment of acute promyelocytic leukemia (APL). ATRA induces both in vitro and in vivo differentiation of APL cells into mature granulocytes. However, the molecular mechanisms involved in ATRA-dependent growth inhibition and cellular differentiation are not presently understood. The NB4 cell line, which is derived from the bone marrow of a patient with APL during relapse, can be used as a model system to study the growth and differentiation of APL cells. Because interferon (IFN) regulatory factors (IRF-1 and IRF-2) and other IFN-inducible gene products regulate cell growth, we analyzed the effects of ATRA on the expression of these genes. We show that ATRA directly activates IRF-1 gene expression, followed by activation of IRF-2 and 2′–5′ oligoadenylate synthetase (OAS) gene expression with slower kinetics. In addition to NB4 cells, ATRA also activated IRF-1 gene expression in HL-60, U937, and THP-1 cells, which all respond to ATRA by growth inhibition. A more than additive increase in IRF-1 gene expression was seen with ATRA and IFN-gamma in NB4 cells. ATRA did not activate nuclear factor kappa B or signal transducer and activator of transcription (STAT) activation pathways, suggesting that an alternate mechanism is involved in IRF-1 gene activation. The ATRA-induced expression of IRF-1, an activator of transcription and repressor of transformation, may be one of the molecular mechanisms of ATRA-induced growth inhibition, and the basis for the synergistic actions of ATRA and IFNs in myeloid leukemia cells.

Abstract 141: Histone Methyltransferase Setdb2 is important for miRNA mediated cardiac reprogramming

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
Imke Kirste ◽  
Tilanthi M Jayawardena ◽  
J. A Payne ◽  
Victor J Dzau ◽  
Maria Mirotsou
Keyword(s):  
Gene Expression ◽  
Cell Fate ◽  
Histone Modifications ◽  
Molecular Mechanisms ◽  
Cardiac Tissue ◽  
Chromatin Modification ◽  
Jak Inhibitor ◽  
Daunting Challenge

Rationale: Regeneration of damaged cardiac tissue after injury presents a daunting challenge in cardiovascular medicine. Recent developments in reprogramming of somatic cells directly to cells of other lineages have raised the possibility of using this approach for cardiac regenerative therapy. Our group recently demonstrated successful miRNA mediated cardiac reprogramming in vitro and in vivo using a combination of miRNAs 1, 133, 206 and 499. Although, the molecular mechanisms underlying miRNA mediated fibroblast reprogramming to cardiomyocytes are yet unknown, accumulating evidence suggest that reprogramming acts through distinct phases and that histone modifications play an important role in these processes. Objective: Identify key genes involved in initiating miRNA mediated reprogramming via histone modifications. Methods and Results: For this, we analyzed the expression levels of 81 different genes involved in chromatin modification 4 days after miRNA transfection using PCR arrays. This analysis revealed that 6 of the 81 tested genes showed differential gene expression (≤-1.5-fold and p <0.02). JAK inhibitor-1 treatment, known for increasing reprogramming efficiency, further enhanced gene expression changes in 5 of these 6 genes. Setdb2, an H3K9 methyltransferase, was one of the most down-regulated targets 4 days after miRNA transfection (-1.4 fold, p<0.001). This effect was enhanced further when miRNAs were combined with the JAK inhibitor-1 (-2.6 fold, p<0.001). Silencing of Setdb2 using siRNAs further accentuated miRNA cardiac reprogramming as measured by cardiac transcription factor expression at 3 days and 6 days post treatment. Similar trends were observed by FACS analysis detecting increased percentage of αMHC-positive cells in siRNA treated fibroblasts compared to control treated only with the miRNA combination. Interestingly, our data showed that Setdb2 silencing alone was sufficient to initiate cardiac reprogramming, suggesting that Setdb2 might play a crucial role in defining cardiac cell fate. Conclusion: In conclusion our results indicate that Setdb2 down-regulation plays an important role in the direct reprogramming of fibroblasts to cardiomyocyte-like cells.

scholarly journals Elevated expression of NEU1 sialidase in idiopathic pulmonary fibrosis provokes pulmonary collagen deposition, lymphocytosis, and fibrosis

2016 ◽  
Vol 310 (10) ◽  
pp. L940-L954 ◽  
Author(s):  
Irina G. Luzina ◽  
Virginia Lockatell ◽  
Sang W. Hyun ◽  
Pavel Kopach ◽  
Phillip H. Kang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Endothelial Cells ◽  
Idiopathic Pulmonary Fibrosis ◽  
Pulmonary Fibrosis ◽  
Molecular Mechanisms ◽  
Transforming Growth Factor ◽  
Recombinant Adenovirus ◽  
Global Gene Expression

Idiopathic pulmonary fibrosis (IPF) poses challenges to understanding its underlying cellular and molecular mechanisms and the development of better therapies. Previous studies suggest a pathophysiological role for neuraminidase 1 (NEU1), an enzyme that removes terminal sialic acid from glycoproteins. We observed increased NEU1 expression in epithelial and endothelial cells, as well as fibroblasts, in the lungs of patients with IPF compared with healthy control lungs. Recombinant adenovirus-mediated gene delivery of NEU1 to cultured primary human cells elicited profound changes in cellular phenotypes. Small airway epithelial cell migration was impaired in wounding assays, whereas, in pulmonary microvascular endothelial cells, NEU1 overexpression strongly impacted global gene expression, increased T cell adhesion to endothelial monolayers, and disrupted endothelial capillary-like tube formation. NEU1 overexpression in fibroblasts provoked increased levels of collagen types I and III, substantial changes in global gene expression, and accelerated degradation of matrix metalloproteinase-14. Intratracheal instillation of NEU1 encoding, but not control adenovirus, induced lymphocyte accumulation in bronchoalveolar lavage samples and lung tissues and elevations of pulmonary transforming growth factor-β and collagen. The lymphocytes were predominantly T cells, with CD8+ cells exceeding CD4+ cells by nearly twofold. These combined data indicate that elevated NEU1 expression alters functional activities of distinct lung cell types in vitro and recapitulates lymphocytic infiltration and collagen accumulation in vivo, consistent with mechanisms implicated in lung fibrosis.

scholarly journals Retinoic acid activates interferon regulatory factor-1 gene expression in myeloid cells

Blood ◽  
1996 ◽  
Vol 88 (1) ◽  
pp. 114-123 ◽  
Author(s):  
S Matikainen ◽  
T Ronni ◽  
M Hurme ◽  
R Pine ◽  
I Julkunen
Keyword(s):  
Gene Expression ◽  
Retinoic Acid ◽  
Growth Inhibition ◽  
Molecular Mechanisms ◽  
Gene Activation ◽  
Oligoadenylate Synthetase ◽  
Nb4 Cells ◽  
Drug Of Choice

Abstract All-trans-retinoic acid (ATRA) is the drug of choice in the treatment of acute promyelocytic leukemia (APL). ATRA induces both in vitro and in vivo differentiation of APL cells into mature granulocytes. However, the molecular mechanisms involved in ATRA-dependent growth inhibition and cellular differentiation are not presently understood. The NB4 cell line, which is derived from the bone marrow of a patient with APL during relapse, can be used as a model system to study the growth and differentiation of APL cells. Because interferon (IFN) regulatory factors (IRF-1 and IRF-2) and other IFN-inducible gene products regulate cell growth, we analyzed the effects of ATRA on the expression of these genes. We show that ATRA directly activates IRF-1 gene expression, followed by activation of IRF-2 and 2′–5′ oligoadenylate synthetase (OAS) gene expression with slower kinetics. In addition to NB4 cells, ATRA also activated IRF-1 gene expression in HL-60, U937, and THP-1 cells, which all respond to ATRA by growth inhibition. A more than additive increase in IRF-1 gene expression was seen with ATRA and IFN-gamma in NB4 cells. ATRA did not activate nuclear factor kappa B or signal transducer and activator of transcription (STAT) activation pathways, suggesting that an alternate mechanism is involved in IRF-1 gene activation. The ATRA-induced expression of IRF-1, an activator of transcription and repressor of transformation, may be one of the molecular mechanisms of ATRA-induced growth inhibition, and the basis for the synergistic actions of ATRA and IFNs in myeloid leukemia cells.

scholarly journals HPV E7-mediated NCAPH ectopic expression regulates the carcinogenesis of cervical carcinoma via PI3K/AKT/SGK pathway

2020 ◽  
Vol 11 (12) ◽  
Author(s):  
Meng Wang ◽  
Xiaowen Qiao ◽  
Tamara Cooper ◽  
Wei Pan ◽  
Liang Liu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cervical Cancer ◽  
Cancer Cells ◽  
Molecular Mechanisms ◽  
Ectopic Expression ◽  
Depth Of Invasion ◽  
Mesenchymal Transition ◽  
Hpv E7

AbstractCervical cancer is one of the most common gynecological tumors in the world, and human papillomavirus (HPV) infection is its causative agent. However, the molecular mechanisms involved in the carcinogenesis of cervical cancer still require clarification. Here we found that knockdown of Non-SMC (Structural Maintenance of Chromosomes) condensin I complex subunit H (NCAPH) gene expression significantly inhibited the proliferation, migration, invasion and epithelial–mesenchymal transition (EMT) of cervical cancer cells in vitro, and restrained xenograft tumor formation in vivo. Intriguingly, HPV E7 could form a positive feedback loop with NCAPH. E7 upregulated NCAPH gene expression via E2F1 which initiated NCAPH transcription by binding to its promoter directly. Silencing of NCAPH reduced E7 transcription via promoting the transition of AP-1 heterodimer from c-Fos/c-Jun to Fra-1/c-Jun. Moreover, the E7-mediated NCAPH overexpression was involved in the activation of the PI3K/AKT/SGK signaling pathway. In vivo, NCAPH expression in cervical cancer tissues was significantly higher than which in normal cervix and high-grade squamous intraepithelial lesion (HSIL) tissues, and its expression was significantly correlated with tumor size, depth of invasion and lymph node metastasis. Patients with high NCAPH expression had a significantly better survival outcomes than those with low-expression, suggesting that NCAPH-induced cell proliferation might sensitize cancer cells to adjuvant therapy. In conclusion, our results revealed the role of NCAPH in the carcinogenesis of cervical cancer in vitro and in vivo. The interaction between E7 and NCAPH expands the mechanism of HPV induced tumorigenesis and that of host genes regulating HPV E7.

A novel colon-specific steroid prodrug enhances sodium chloride absorption in rat colitis

1995 ◽  
Vol 269 (2) ◽  
pp. G210-G218
Author(s):  
R. N. Fedorak ◽  
N. Cui ◽  
D. R. Friend ◽  
K. L. Madsen ◽  
L. R. Empey
Keyword(s):  
Sodium Chloride ◽  
Colonic Mucosa ◽  
Epithelial Transport ◽  
Free Drug ◽  
Fluid Absorption ◽  
Ussing Chambers ◽  
Mucosal Integrity ◽  
Chloride Absorption

A recently synthesized novel colon-specific dexamethasone prodrug, dexamethasone-beta-D-glucuronide, delivers efficacious amounts of dexamethasone to the colon with limited adrenal suppressive effects. During experimentally induced colitis in rats, the dexamethasone prodrug is significantly more potent than free dexamethasone in improving colonic fluid and electrolyte absorptive injury. The present studies examined whether the improvement in colonic absorption seen with the prodrug occurred as a consequence of alterations in sodium and chloride epithelial transport. The efficacy of the dexamethasone prodrug and free dexamethasone were tested in an acetic acid-induced rat model of colitis. Healing of the induced colitis was assessed by measuring net colonic fluid absorption and surface area ulceration. Transmural unidirectional fluxes of 22Na and 36Cl across sheets of colonic mucosa were measured in Ussing chambers. Treatment of colitis with the prodrug delivered a sixfold higher concentration of dexamethasone to the colon than did treatment with the free drug. The prodrug accelerated healing of colitis by returning in vivo colonic fluid absorption to normal and virtually eliminated colonic macroscopic ulceration, whereas the free drug did not. In vitro transmural fluxes demonstrated that, in addition to repair of mucosal integrity, the prodrug enhanced electroneutral sodium chloride absorption over and above that seen in control animals or after treatment with the free drug. Both the prodrug and the free drug limited theophylline-mediated net chloride and sodium secretion, an effect that would be consistent with the antidiarrheal effect induced by these drugs in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Involvement of Histone Lysine Methylation in p21 Gene Expression in Rat KidneyIn Vivoand Rat Mesangial CellsIn Vitrounder Diabetic Conditions

2016 ◽  
Vol 2016 ◽  
pp. 1-12 ◽  
Author(s):  
Xiangjun Li ◽  
Chaoyuan Li ◽  
Xiaoxia Li ◽  
Peihe Cui ◽  
Qifeng Li ◽  
...  
Keyword(s):  
Gene Expression ◽  
Molecular Mechanisms ◽  
Mesangial Cells ◽  
Histone H3 ◽  
Diabetic Rats ◽  
Lysine Methylation ◽  
Histone Lysine Methylation ◽  
Histone Lysine

Diabetic nephropathy (DN), a common complication associated with type 1 and type 2 diabetes mellitus (DM), characterized by glomerular mesangial expansion, inflammation, accumulation of extracellular matrix (ECM) protein, and hypertrophy, is the major cause of end-stage renal disease (ESRD). Increasing evidence suggested that p21-dependent glomerular and mesangial cell (MC) hypertrophy play key roles in the pathogenesis of DN. Recently, posttranscriptional modifications (PTMs) have uncovered novel molecular mechanisms involved in DN. However, precise regulatory mechanism of histone lysine methylation (HKme) mediating p21 related hypertrophy associated with DN is not clear. We evaluated the roles of HKme and histone methyltransferase (HMT) SET7/9 in p21 gene expression in glomeruli of diabetic rats and in high glucose- (HG-) treated rat mesangial cells (RMCs). p21 gene expression was upregulated in diabetic rats glomeruli; chromatin immunoprecipitation (ChIP) assays showed decreased histone H3-lysine9-dimethylation (H3K9me2) accompanied with enhanced histone H3-lysine4-methylation (H3K4me1/3) and SET7/9 occupancies at the p21 promoter. HG-treated RMCs exhibited increased p21 mRNA, H3K4me level, SET7/9 recruitment, and inverse H3K9me, which were reversed by TGF-β1 antibody. These data uncovered key roles of H3Kme and SET7/9 responsible for p21 gene expressionin vivoandin vitrounder diabetic conditions and confirmed preventive effect of TGF-β1 antibody on DN.

scholarly journals Analysis of the Mouse CSF-1 Gene Promoter in a Transgenic Mouse Model1

2003 ◽  
Vol 51 (7) ◽  
pp. 941-949 ◽  
Author(s):  
Sherry L. Abboud ◽  
Maria Bunegin ◽  
Nandini Ghosh-Choudhury ◽  
Kathleen Woodruff
Keyword(s):  
Gene Expression ◽  
Transgenic Mice ◽  
Transgene Expression ◽  
Transcriptional Activity ◽  
Molecular Mechanisms ◽  
Tissue Expression ◽  
Cellular Level ◽  
Liver And Kidney

CSF-1 stimulates monocyte and osteoclast populations. However, the molecular mechanisms involved in regulating CSF-1 gene expression are unclear. To identify regulatory regions that control normal CSF-1 gene expression, a −774/+183-bp fragment of the murine CSF-1 promoter was analyzed in vitro and in vivo. Transcriptional activity was high in cultured osteoblasts that express CSF-1 mRNA compared to ARH-77 B cells that lack CSF-1 gene expression. Transient transfection of osteoblasts with promoter deletion constructs showed that the −774-bp fragment conferred the highest transcriptional activity and contained activator and repressor sequences. To assess the ability of the CSF-1 promoter to confer normal tissue expression of CSF-1, transgenic mice containing the −774/+183-bp region driving the E. coli β-galactosidase (lacZ) reporter gene were generated. β-Gal analysis of whole tissue extracts showed transgene expression in all tissues tested except liver and kidney. At the cellular level, the pattern of β-gal expression in the spleen, thymus, bone, lung, and testes of adult transgenic mice mimicked normal endogenous CSF-1 mRNA expression in non-transgenic littermates detected by in situ hybridization. This region also directed appropriate transgene expression to sites in other tissues known to synthesize CSF-1, with the exception of the liver and kidney. These findings indicate that the −774-bp fragment contains cis-acting elements sufficient to direct CSF-1 gene expression in many tissues. CSF-1 promoter/lacZ mice may be useful for studying the transcriptional mechanisms involved in regulating CSF-1 gene expression in tissues throughout development.

scholarly journals CHIP Represses Myocardin-Induced Smooth Muscle Cell Differentiation via Ubiquitin-Mediated Proteasomal Degradation

2009 ◽  
Vol 29 (9) ◽  
pp. 2398-2408 ◽  
Author(s):  
Ping Xie ◽  
Yongna Fan ◽  
Hua Zhang ◽  
Yuan Zhang ◽  
Mingpeng She ◽  
...  
Keyword(s):  
Gene Expression ◽  
Smooth Muscle ◽  
Molecular Mechanisms ◽  
Serum Response Factor ◽  
Ex Vivo ◽  
Critical Role ◽  
Physiological Control ◽  
Interacting Protein

ABSTRACT Myocardin, a coactivator of serum response factor (SRF), plays a critical role in the differentiation of vascular smooth muscle cells (SMCs). However, the molecular mechanisms regulating myocardin stability and activity are not well defined. Here we show that the E3 ligase C terminus of Hsc70-interacting protein (CHIP) represses myocardin-dependent SMC gene expression and transcriptional activity. CHIP interacts with and promotes myocardin ubiquitin-mediated degradation by the proteasome in vivo and in vitro. Furthermore, myocardin ubiquitination by CHIP requires its phosphorylation. Importantly, CHIP overexpression reduces the level of myocardin-dependent SMC contractile gene expression and diminishes arterial contractility ex vivo. These findings for the first time, to our knowledge, demonstrate that CHIP-promoted proteolysis of myocardin plays a key role in the physiological control of SMC phenotype and vessel tone, which may have an important implication for pathophysiological conditions such as atherosclerosis, hypertension, and Alzheimer's disease.

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