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At least 883 records · Page 49Linked to original sources
PubMed1996

Transgenic rice plants harboring an introduced potato proteinase inhibitor II gene are insect resistant.

We introduced the potato proteinase inhibitor II (PINII) gene (pin2) into several Japonica rice varieties, and regenerated a large number of transgenic rice plants. Wound-inducible expression of the pin2 gene driven by its own promoter, together with the first intron of the rice actin 1 gene (act1), resulted in high-level accumulation of the PINII protein in the transgenic plants. The introduced pin2 gene was stably inherited in the second, third, and fourth generations, as shown by molecular analyses. Based on data from the molecular analyses, several homozygous transgenic lines were obtained. Bioassay for insect resistance with the fifth-generation transgenic rice plants showed that transgenic rice plants had increased resistance to a major rice insect pest, pink stem borer (Sesamia inferens). Thus, introduction of an insecticidal proteinase inhibitor gene into cereal plants can be used as a general strategy for control of insect pests.

Animals↗
PubMed1996

Constitutive class I-restricted exogenous presentation of self antigens in vivo.

Ovalbumin (OVA)-specific CD8+ T cells from the T cell receptor-transgenic line OT-I (OT-I cells) were injected into unirradiated transgenic RIP-mOVA mice, which express a membrane-bound form of OVA (mOVA) in the pancreatic islet beta cells and the renal proximal tubular cells. OT-I cells accumulated in the draining lymph nodes (LN) of the kidneys and pancreas and in no other LN. They displayed an activated phenotype and a proportion entered cell cycle. Unilateral nephrectomy 7-13 d before inoculation of OT-I cells into RIP-mOVA mice allowed the injected T cells to home only to the regional LN of the remaining kidney (and pancreas), but when the operation was performed 4 h before injecting the T cells, homing to the LN of the excised kidney was evident. When the bone marrow of RIP-mOVA mice was replaced with one of a major histocompatibility haplotype incapable of presenting OVA to OT-I cells, no homing or activation was detectable. Therefore, OT-I cells were activated by OVA presented by short-lived antigen-presenting cells of bone marrow origin present in the draining LN of OVA-expressing tissue. These results provide the first evidence that tissue-associated "self" antigens can be presented in the context of class I via an exogenous processing pathway. This offers a constitutive mechanism whereby T cells can be primed to antigens that are present in nonlymphoid tissues, which are not normally surveyed by recirculating naive T cells.

Animals↗
PubMed1997

Genetic modification of the phenotypes produced by amyloid precursor protein overexpression in transgenic mice.

Overexpression of Alzheimer amyloid precursor protein (APP) produces dramatically different phenotypes in transgenic mice depending on the genetic background. For example, concentrations of APP that produce amyloid plaques in outbred transgenic lines are lethal for inbred FVB/N or C57BL/6J mice. Expression of SOD1 transgenes is protective, suggesting involvement of oxidative damage in premature death, but ablation of Apoe had no significant effect. In contrast, FGF2 transgene overexpression enhances the lethal effects of APP. Differential survival does not appear to reflect genetic differences in APP processing, but rather host responses to APP or its derivatives.

Alleles↗
PubMed2006

Sequence elements critical for efficient RNA editing of a tobacco chloroplast transcript in vivo and in vitro.

In tobacco chloroplast transcripts 34 nt are efficiently edited to U. No common consensus region is present around all editing sites; however, sites can be grouped in clusters that share short common sequences. Transgene transcripts carrying either the wild-type -31/+22 or -31/+60 sequence near NTrpoB C473, an editing site within tobacco rpoB transcripts, or three different mutated sequences, were all highly edited in vivo. Endogenous transcripts of rpoB, psbL and rps14, all of which contain common sequences S1, S2 and S3 5' to NTrpoB C473, NTpsbL C2 and NTrps14 C80, were less edited in transgenic plants that over-express transcripts from NTrpoB C473 transgenes. Extent of reduction of endogenous editing differed between transgenic lines expressing mutated -31/+22 regions, depending on the abundance of the transgene transcripts. The -20/-5 sequence contains critical 5' sequence elements. Synthetic RNA templates with alterations within this 5' region were less efficiently edited in vitro than wild-type templates, by either tobacco or maize chloroplast extracts. The tobacco chloroplast extract supports both RNA editing and processing of 3' transcript termini. We conclude that within the -20/-5 region, sequences common to editing sites in the transcripts of rpoB, psbL and rps14 are critical for efficient NTrpoB C473 editing.

Base Sequence↗
PubMedDate not supplied

Overexpression of a calcium-binding protein, S100 beta, in astrocytes alters synaptic plasticity and impairs spatial learning in transgenic mice.

Recent evidence suggests that slowly propagating Ca2+ waves from astrocytes can modulate the function of neurons. Altering astrocytic calcium processes in vivo may therefore affect neuronal and behavioral phenotypes. Previously, we generated transgenic mice that overexpress an astrocytic calcium-binding protein, S100 beta. Immunocytochemistry and in situ hybridization showed elevated expression in the astrocytes of the hippocampus and other brain regions. Neurons in the hippocampus were negative for S100 beta. In this paper we analyze the hippocampal electrophysiology and learning properties of mice from two transgenic lines. Significant differences were found between the hippocampal slices of normal and transgenic mice in their response to high frequency (100 Hz) stimulation. The overall distribution of post-tetanic excitatory postsynaptic potentials (EPSP) of the slices from the transgenic mice was shifted significantly toward smaller values to a degree that 25% of slices exhibited depression. The altered hippocampal neurophysiology was accompanied by an impairment in a hippocampal-dependent learning task. Transgenic mice showed significant impairment in a spatial version of the Morris water maze, however, they performed normally in non-spatial tasks. Probe trials showed that transgenic mice, though significantly impaired, also acquired spatial information. The results suggested that the impairment was not due to motor dysfunction, impaired vision or motivation of the transgenic mice, findings compatible with a possible hippocampal mechanism. We conclude that overexpression of S100 beta in astrocytes impairs, but does not abolish, the ability to solve a spatial task, and it leads to a significantly decreased post-tetanic potentiation in the hippocampal slice. We hypothesize that the changes are due to calcium mediated processes. Our results support the notion that astrocytes are involved in higher brain functions.

Animals↗
PubMed2006

Analysis of cell migration, transdifferentiation and apoptosis during mouse secondary palate fusion.

Malformations in secondary palate fusion will lead to cleft palate, a common human birth defect. Palate fusion involves the formation and subsequent degeneration of the medial edge epithelial seam. The cellular mechanisms underlying seam degeneration have been a major focus in the study of palatogenesis. Three mechanisms have been proposed for seam degeneration: lateral migration of medial edge epithelial cells; epithelial-mesenchymal trans-differentiation; and apoptosis of medial edge epithelial cells. However, there is still a great deal of controversy over these proposed mechanisms. In this study, we established a [Rosa26<-->C57BL/6] chimeric culture system, in which a Rosa26-originated ;blue' palatal shelf was paired with a C57BL/6-derived ;white' palatal shelf. Using this organ culture system, we observed the migration of medial edge epithelial cells to the nasal side, but not to the oral side. We also observed an anteroposterior migration of medial edge epithelial cells, which may play an important role in posterior palate fusion. To examine epithelial-mesenchymal transdifferentiation during palate fusion, we bred a cytokeratin 14-Cre transgenic line into the R26R background. In situ hybridization showed that the Cre transgene is expressed exclusively in the epithelium. However, beta-galactosidase staining gave extensive signals in the palatal mesenchymal region during and after palate fusion, demonstrating the occurrence of an epithelial-mesenchymal transdifferentiation mechanism during palate fusion. Finally, we showed that Apaf1 mutant mouse embryos are able to complete palate fusion without DNA fragmentation-mediated programmed cell death, indicating that this is not essential for palate fusion in vivo.

Animals↗
PubMed2005

Production of transgenic models in hypertension.

This chapter describes the generation of transgenic mice and rats by microinjection of DNA constructs into the pronucleus of a zygote. The transgene DNA is randomly integrated as several tandem copies at one site into the genome and is transmitted to the offspring of the founder animal derived from the injected embryo, thereby creating a stable transgenic line. The technology includes the following steps: design and generation of the transgene construct, superovulation of donor animals, isolation of fertilized oocytes, microinjection of the transgene construct into one pronucleus of the zygotes, transfer of injected embryos into the oviduct of a foster mother, and identification of transgenic animals in the offspring.

Animals↗
PubMed1997

Viability of transgenic rat embryos after freezing and thawing.

In-vivo viability of frozen-thawed embryos derived from transgenic rats, as well as the transmission and the expression of transgenes in the resultant newborn rats, was investigated. Three strains of transgenic rats, carrying human growth hormone gene connected downstream to the promoter region of the bovine alpha-lactalbumin gene (alpha LA/hGH), bovine beta-casein gene (beta CN/hGH) or bovine alpha-S1 casein gene (alpha S1CN/hGH), were used. Two-cell stage embryos (non-transgenic Wistar female x heterozygous transgenic male) were placed in 10% (v/v) dimethylsulfoxide (DMSO) solution and cooled from -7 to -30 degrees C at -0.5 degree C/min before being plunged into liquid nitrogen. After 2 to 4 years storage, the embryos were thawed by rapid warming. The intact embryos were transferred into the oviducts of Day 1 pseudopregnant recipients. The postthaw survival rate of frozen embryos was high in all 3 transgenic strains (88 to 92%), which was similar to that of control (non-transgenic) frozen embryos (95%). Development to newborn rats following transfer of embryos derived from the 3 strains (64 to 68%) was also similar to that of control embryos (60%). These transgenes (alpha LA/hGH, beta CN/hGH and alpha S1CN/hGH) were detected in the DNA extracts from tail tissue of the newborn rats, but the transmission rates (41, 23 and 32%, respectively) were lower than 50% which is expected in the Mendelian fashion. In a transgenic line carrying alpha S1CN/hGH, hGH levels of secretion into the milk of transgenic newborn rats derived from frozen-thawed embryos and her transgenic offspring were the same mg/ml-level as that of their founder rat. Two-step freezing of embryos derived from transgenic rats was therefore an effective method for the long-term cryopreservation of transgene.

Animals↗
PubMed2003

Transgenic expression of CD36 in the spontaneously hypertensive rat is associated with amelioration of metabolic disturbances but has no effect on hypertension.

Spontaneously hypertensive rats (SHR/NIH strain) harbor a deletion variant in the Cd36 fatty acid transporter and display defective fatty acid metabolism, insulin resistance and hypertension. Transgenic rescue of Cd36 in SHR ameliorates insulin resistance and improves dyslipidemia. However, the role of Cd36 in blood pressure regulation remains controversial due to inconsistent blood pressure effects that were observed with transgenic expression of Cd36 on the SHR background. In the current studies, we developed two new SHR transgenic lines, which express wild type Cd36 under the control of the universal Ef-1 alpha promoter, and examined the effects of transgenic expression of wild type Cd36 on selected metabolic and cardiovascular phenotypes. Transgenic expression of Cd36 in the new lines was associated with significantly decreased serum fatty acids, amelioration of insulin resistance and glucose intolerance but failed to induce any consistent changes in blood pressure as measured by radiotelemetry. The current findings confirm the genetic association of defective Cd36 with disordered insulin action and fatty acid metabolism in the SHR/NIH strain and suggest that Cd36 is linked to other gene(s) on rat chromosome 4 that regulate blood pressure.

Adipose Tissue↗
PubMed2001

hGFAP-cre transgenic mice for manipulation of glial and neuronal function in vivo.

With the goal of performing astrocyte-specific modification of genes in the mouse, we have generated a transgenic line expressing Cre recombinase under the control of the human glial fibrillary acidic protein (hGFAP) promoter. Activity was monitored by crossing the hGFAP-cre transgenics with either of two reporter lines carrying a lacZ gene whose expression requires excision of loxP-flanked stop sequences. We found that lacZ expression was primarily limited to the central nervous system, but therein was widespread in neurons and ependyma. Cell types within the brain that notably failed to activate lacZ expression included Purkinje neurons of the cerebellum and choroid plexus epithelium. Onset of Cre expression began in the forebrain by e13.5, suggesting that the hGFAP promoter is active in a multi-potential neural stem cell.

Animals↗
PubMed2005

Use of tissue recombination to predict phenotypes of transgenic mouse models of prostate carcinoma.

Transgenic mouse models of cancer represent a powerful approach for exploring disease processes and testing potential therapeutic interventions. Currently, it is difficult to predict if a specific genetic manipulation will result in a desirable phenotype. The present study tests the idea that tissue recombinants recapitulate the pathologic features of the neoplastic prostate seen in transgenic mice, and would thus be suitable predictive models for new mouse design. The large probasin-large T-antigen (LPB-Tag) transgenic lines 12T-7f and 12T-10 were used as a basis for this study. Tissue recombinants of bladder epithelium (BlE) and urogenital sinus mesenchyme (UGM) were implanted under the renal capsule of athymic mice. Recombinants composed of BlE from 12T-10 LPB-Tag and wild-type (wt) UGM faithfully recapitulated the histopathologic and temporal features of intact transgenic mice of this line. Tissue recombinants using BlE from 12T-7f mice and wt UGM developed epithelial proliferation with atypia that lacked the associated hypercellular stroma seen in the intact 12T-7f line. Recombinants using 12T-7f UGM demonstrated that the hypercellular stroma results from stromal cell expression of the SV40 large T antigen. Corresponding to the recombinant phenotypes, stromal Tag immunostaining was observed in prostate tissues from intact 12T-7f but not 12T-10 mice. Similar stromal expression of Tag was also noted in the hypercellular TRAMP prostatic stroma. Further analysis revealed a previously unreported pattern of SV40T expression in the LADY and TRAMP models including ductus deferens and seminal vesicle stroma as well as region and cell type-specific patterns in the epididymis. The present study demonstrates the utility of using tissue recombination to explore organ-specific phenotypes. Recombination strategies should enable quick and cost-effective screening for likely phenotypes in transgenic animals. This comparison of tissue recombination to existing models shows that this approach can elicit new information on well-characterized models.

Animals↗
PubMed2003

A transgenic insect cell line engineered to produce CMP-sialic acid and sialylated glycoproteins.

We have previously engineered transgenic insect cell lines to express mammalian glycosyltransferases and showed that these cells can sialylate N-glycoproteins, despite the fact that they have little intracellular sialic acid and no detectable CMP-sialic acid. In the accompanying study, we presented evidence that these cell lines can salvage sialic acids for de novo glycoprotein sialylation from extracellular sialoglycoproteins, such as fetuin, found in fetal bovine serum. This finding led us to create a new transgenic insect cell line designed to synthesize its own sialic acid and CMP-sialic acid. SfSWT-1 cells, which encode five mammalian glycosyltransferases, were transformed with two additional mammalian genes that encode sialic acid synthase and CMP-sialic acid synthetase. The resulting cell line expressed all seven mammalian genes, produced CMP-sialic acid, and sialylated a recombinant glycoprotein when cultured in a serum-free growth medium supplemented with N-acetylmannosamine. Thus the addition of mammalian genes encoding two enzymes involved in CMP-sialic acid biosynthesis yielded a new transgenic insect cell line, SfSWT-3, that can sialylate recombinant glycoproteins in the absence of fetal bovine serum. This new cell line will be widely useful as an improved host for baculovirus-mediated recombinant glycoprotein production.

Animals↗
PubMed1996

Expression in brain of amyloid precursor protein mutated in the alpha-secretase site causes disturbed behavior, neuronal degeneration and premature death in transgenic mice.

A double mutation in the alpha-secretase site in the betaA4 region of mouse amyloid precursor protein (APP) reduced its secretion from COS cells, polarized MDCK cells and rat primary neurons. Expression of this mutant in the brain of mice, using the neuron-specific elements of the mouse Thy-1 gene promoter, resulted in transgenic mice that became progressively hyperactive, displayed seizures and died prematurely. In three different transgenic lines the severity of the phenotype was related directly to the expression levels of the transgene, estimated by both mRNA and protein levels. In addition, homozygous mice derived from each transgenic strain showed more severe symptoms which also occurred earlier in life than in heterozygotes. The observed symptoms were, however, not essentially different in the different lines. Increased aggressiveness, disturbed responses to kainic acid and N-methyl-D-aspartate, neophobia and deficiency in exploratory behavior were demonstrated in these mice. In the brain, the observed neuropathological changes included necrosis, apoptosis and astrogliosis in the hippocampus, cortex and other areas. The data demonstrate that incomplete or incorrect alpha-secretase processing of APP results in severe neurotoxicity and that this effect is expressed in a dominant manner.

Amyloid Precursor Protein Secretases↗
PubMed1999

Transgenic rats expressing the human ET-2 gene: a model for the study of endothelin actions in vivo.

Previous studies have characterized the endothelin peptides (ET-1, ET-2, ET-3) as strong vasoconstrictors which are possibly involved in the pathogenesis of cardiovascular disease. Whereas ET-1 and ET-3 have been characterized using a number of approaches, little is known about the function of ET-2. The aim of this study was to define the role of ET-2 in physiology and pathophysiology using a transgenic approach. Transgenic rats expressing a genomic construct of the human ET-2 gene were generated by microinjection of fertilized oocytes from Sprague-Dawley rats. Two transgenic lines were generated, and one line was further characterized in detail. Studies on mRNA expression demonstrated that the transgene is expressed predominantly in kidney, gastrointestinal tract, adrenal gland, lung, and brain. Plasma endothelin levels were elevated 2-fold, and big-endothelin levels were elevated 2.5-fold. Despite these alterations blood pressure in transgenic rats remained normal. Further analysis of transgenic animals revealed that endothelin receptors were not downregulated, and that infusion of exogenous human ET-2 results in an enhanced blood pressure response. These observations suggest the presence of counterregulatory mechanisms influencing the effects of endothelin on blood pressure. One of these mechanisms may involve the nitric oxide system since infusion of an inhibitor of nitric oxide synthase resulted in a greater blood pressure response than in non-transgenic littermates. Despite unchanged blood pressure, alterations were observed in organ development and function, namely of hearts and kidneys, indicating an interference between transgene expression and growth processes. Male rats seem to be more susceptible to endothelin actions. These data show that the elevation in endothelin-2 expression in this transgenic model does not induce hypertension but leads to changes at the end-organ level. Normotension is most likely due to compensatory mechanisms such as increased nitric oxide formation.

Animals↗
PubMed2001

Erythrocyte ankyrin promoter mutations associated with recessive hereditary spherocytosis cause significant abnormalities in ankyrin expression.

Ankyrin defects are the most common cause of hereditary spherocytosis (HS). In several kindreds with recessive, ankyrin-deficient HS, mutations have been identified in the ankyrin promoter that have been proposed to decrease ankyrin synthesis. We analyzed the effects of two mutations, -108T to C and -108T to C in cis with -153G to A, on ankyrin expression. No difference between wild type and mutant promoters was demonstrated in transfection or gel shift assays in vitro. Transgenic mice with a wild type ankyrin promoter linked to a human (A)gamma-globin gene expressed gamma-globin in 100% of erythrocytes in a copy number-dependent, position-independent manner. Transgenic mice with the mutant -108 promoter demonstrated variegated gamma-globin expression, but showed copy number-dependent and position-independent expression similar to wild type. Severe effects in ankyrin expression were seen in mice with the linked -108/-153 mutations. Three transgenic lines had undetectable levels of (A)gamma-globin mRNA, indicating position-dependent expression, and four lines expressed significantly lower levels of (A)gamma-globin mRNA than wild type. Two of four expressing lines showed variegated gamma-globin expression, and there was no correlation between transgene copy number and RNA level, indicating copy number-independent expression. These data are the first demonstration of functional defects caused by HS-related, ankyrin gene promoter mutations.

Animals↗
PubMed2005

Molecular mechanisms of hepatocarcinogenesis in transgenic mouse models of liver cancer.

Overexpression of c-myc and transforming growth factor-alpha (TGF-alpha) has been frequently observed in human hepatocellular carcinoma (HCC),suggesting a pivotal role played by these protooncogenes in liver oncogenesis. In order to investigate the molecular events underlying human hepatic malignant transformation, we have generated c-myc and c-myc/ TGF-alpha transgenic mice that are prone to liver cancer. These transgenic mice develop HCCs with different incidence, kinetics and histopathological features. Indeed, co-expression of c-myc and TGF-alpha transgenes results in a dramatic synergistic effect on liver tumor development when compared with respective single transgenic lines, including a shorter latency period and a more aggressive phenotype. The more malignant histopathological features characteristic of c-myc/ TGF-alpha HCCs are the result of the increased proliferation and reduced apoptosis in this model of liver cancer when compared with single parental lines. Accordingly, c-myc and c-myc/l TGF-alpha transgenic mice display a different molecular pathogenesis of HCC. Importantly, the genetic and molecular mechanisms that are involved in c-myc and c-myc/ TGF-alpha liver cancer development are major oncogenic events in human hepatocarcinogenesis, indicating that these mouse models represent a useful tool to dissect and elucidate the molecular basis of human HCC.

Animals↗
PubMed2000

Local production of anti-CD4 antibody by transgenic allogeneic grafts affords partial protection.

BACKGROUND: Immunosuppressive drugs and anti-lymphocyte antibody are used clinically to suppress cellular rejection responses. However, these systemic regimens often led to general immunodeficiency and thus increased susceptibility to opportunistic infection and neoplasia. Immunosuppressive molecules delivered locally may be a way of inhibiting rejection responses, whereas systemic immunity is preserved. To achieve protective local immunosuppression, we produced a graft secreting its own immunomodulator, by deriving transgenic mice expressing a chimeric anti-CD4 antibody (GK2c) in the pancreas. METHODS AND RESULTS: Transgenic mice in bml genetic background expressing a modified anti-mouse CD4 antibody (GK2c) under two promoters have been produced. Tissue expression of GK2c was detected by immunoperoxidase staining. Under the cytomegalovirus promoter, there was abundant GK2c expression in pancreatic exocrine tissue. Under the rat preproinsulin II promoter, there was abundant GK2c expression in pancreatic endocrine tissue only. High-expression transgenic lines had 10-100 microg/ml GK2c in blood plasma. By flow cytometry, these transgenic mice were devoid of CD4+ cells in their peripheral lymphoid organs. To test transgenic mice as donors, fetal pancreata from transgenic mice were grafted into fully allogeneic CBA mice under the kidney capsule, transgenic grafts had prolonged survival compared with control non-transgenic grafts. Furthermore, GK2c transgenic grafts had reduced infiltration with an absence of CD4+ cells at the graft site without any effect on the cell composition in lymphatic tissues. CONCLUSION: Transgenic grafts that secrete anti-CD4 antibody can afford some protection against graft rejection, while only affecting the CD4 population at the graft site.

Animals↗