PubMed Health⌕ Search

Biomedical subjects

J E Wilkinson

Publications and source records attributed to J E Wilkinson.

At least 19 recordsLinked to original sources
PubMed1997

Combined effects of insulin treatment and adipose tissue-specific agouti expression on the development of obesity.

The agouti gene product is a secreted protein that acts in a paracrine manner to regulate coat color in mammals. Several dominant mutations at the agouti locus in mice cause the ectopic, ubiquitous expression of agouti, resulting in a condition similar to adult-onset obesity and non-insulin-dependent diabetes mellitus. The human agouti protein is 85% homologous to mouse agouti; however, unlike the mouse agouti gene, human agouti is normally expressed in adipose tissue. To address whether expression of agouti in human adipose tissue is physiologically relevant, transgenic mice were generated that express agouti in adipose tissue. Similar to most humans, these mice do not become obese or diabetic. However, we found that daily insulin injections significantly increased weight gain in the transgenic lines expressing agouti in adipose tissue, but not in nontransgenic mice. These results suggest that insulin triggers the onset of obesity and that agouti expression in adipose tissue potentiates this effect. Accordingly, the investigation of agouti's role in obesity and non-insulin-dependent diabetes mellitus in mice holds significant promise for understanding the pathophysiology of human obesity.

Adipose Tissue↗
PubMed1997

Isolation and characterization of liver epithelial cell lines from wild-type and mutant TgN737Rpw mice.

The Tg737 gene encodes a tetratricopeptide repeat containing protein that, when disrupted in TgN737Rpw mutant mice, results in pleiotropic phenotypes that include the proliferation of epithelial cells. In the kidney and liver, this causes a phenotype that resembles autosomal recessive polycystic kidney disease. In the liver, the affected epithelial cells morphologically and immunologically resemble oval cells. Here we describe the isolation, culture, and characterization of epithelial cell lines derived from the livers of wild-type, heterozygous, and homozygous TgN737Rpw mice. Essentially homogeneous cell cultures were established and the expression of liver markers was examined by reverse transcriptase polymerase chain reaction and by immunohistochemistry. All of the cell lines reacted to the A6 antibody that was raised against mouse oval cells and expressed markers seen in oval cells. Cells transplanted into the interscapular fat pads of isogenic mice formed well defined ductular structures. Furthermore, in transfection experiments, we have demonstrated the involvement of Tg737 in cellular proliferation.

Adipose Tissue↗
PubMed1996

Functional correction of renal defects in a mouse model for ARPKD through expression of the cloned wild-type Tg737 cDNA.

Autosomal recessive polycystic kidney disease (ARPKD) is characterized by the formation of large collecting tubule and ductular cysts that often result in renal insufficiency within the first decade of life. Understanding the process leading to cyst formation will require the identification and characterization of genes involved in the etiology of this disease. In this regard, we previously described the generation of a mouse model (TgN737Rpw) for ARPKD and the cloning of a candidate gene. Here we show direct involvement of the Tg737 gene in collecting duct cyst formation by expressing the wild-type Tg737 cDNA as a transgene in TgN737Rpw mutants. In contrast to TgN737Rpw mutants, the "rescued" animals survive longer, have normal renal function and normal localization of the EGFr to the basolateral surfaces of collecting duct epithelium.

Animals↗
PubMed1996

Ovine endometrial expression of transforming growth factor beta isoforms during the peri-implantation period.

During the estrous cycle and early pregnancy, the uterus undergoes a variety o morphological changes. Because of their powerful effects on angiogenesis, on extracellular matrix modification, and on cellular proliferation and differentiation, the transforming growth factor beta (TGF beta) family of polypeptides may be involved in the regulation of pregnancy-related endometrial modification. In this study, endometrial stead-state mRNA expression of the three isoforms, TGF beta 1, beta 2, and beta 3, was quantified, and the proteins were localized during the later part of the estrous cycle (Day 13 and 16) and during early pregnancy (Days 13 through 30) in sheep. TGF beta 1 mRNA was expressed as a single transcript with steady-state mRNA expression levels 2-fold higher on Day 16 of the estrous cycle than on Day 13 of the estrous cycle (p < or = 0.002) or Days 13 and 16 (p < or = 0.004) and 0.008, respectively) of gestation. During pregnancy, levels increased progressively to peak on Day 27 (2.5-fold above Day 16 of pregnancy; p < or = 0.0001) and then leveled off at Day 30. Immunocytochemical localization of TGF beta 1 demonstrated protein in glandular protein in glandular and luminal epithelium at all days examined. TGF beta 2 mRNA was expressed as five distinct transcripts, and mRNA expression levels were lowest in Day 16 pregnant endometrium. Thereafter the expression level increased steadily through Day 30 (p < or = 0.0003). TGF beta 2 protein was localized in epithelium, diffusely within the endometrial stroma, and in leukocyte-like cells within the stroma. TGF beta 3 was expressed as one major transcripts and two minor transcripts. As with TGF beta 1, there was a dramatic difference in TGF beta 3 levels between Day 16 of pregnancy and Day 16 of the estrous cycle (3.8-fold, p < or = 0.0001). In pregnant ewes, endometrial TGF beta 3 levels increased 1.9-fold (p = 0.13) between Days 16 and 23 and remained relatively constant through Day 30. Immunohistochemistry localized TGF beta 3 protein most prominantly in the subepithelial stroma of the caruncule from Day 16 of the cycle in endometrial tissue. The observed changes in mRNA and protein expression patterns of TGF beta s within the ovine endometrium suggest that TGF beta s play a role in restructuring and modifying endometrium for a subsequent estrous cycle and/or pregnancy.

Animals↗
PubMed1996

Transforming growth factor beta in bovine placentas.

Transforming growth factor beta s (TGF beta) are a family of multifunctional growth factors that are important embryonic morphogens. Because TGF beta s may regulate the development of epitheliochorial placentas, we investigated the location, expression, secretion, and effects of TGF beta s in bovine placentomes and cell cultures derived from chorionic and endometrial epithelia. Placentomes from early second-trimester pregnancies were examined by immunohistochemistry for TGF beta 1, TGF beta 2, and TGF beta 3, and for TGF beta expression in Northern slot-blots. Effects of TGF beta s were assessed in trophoblastic and endometrial epithelial cell lines by DNA synthesis assays. Secretion of TGF beta s by trophoblastic and endometrial epithelial cells was determined using bioassays. All forms of TGF beta were immunolocalized in bovine placentomes. TGF beta mRNA was expressed in chorioallantois, caruncles, and in cultured trophoblastic and endometrial epithelial cells. Endometrial and trophoblastic cells secreted active and latent TGF beta s, and these cells had a transient proliferative response to all forms of TGF beta. These results indicate that TGF beta s are present at the fetal-maternal interface of the bovine placentome and may promote endometrial and chorionic growth.

Animals↗
PubMed1996

Oval cell proliferation associated with the murine insertional mutation TgN737Rpw.

The Tg737 gene was identified by its direct association with a transgene-induced insertion mutation in the mouse. This mutation causes pleiotropic phenotypes including a syndrome similar to autosomal recessive polycystic kidney disease in humans. This syndrome, in addition to renal cyst formation, includes the presence of an invariably associated liver abnormality. The liver pathology in TgN737Rpw mice is characterized by a biliary hyperplasia that includes the proliferation of cells that morphologically and immunologically resemble oval cells, a liver progenitor cell. This abnormality is first observed at approximately 5 days of age in the portal region and then progresses into the periportal regions. Additionally, the formation and proliferation of dysplastic ductular structures are observed from the onset of the phenotype. Serum chemistry indicated that the primary defect is likely to be of biliary origin, and hepatic function appears normal in the mutant mice. Therefore, this mutation is unlike other causes of oval cell proliferation in that the hepatic parenchyma is relatively unaffected. The identification of the Tg737 gene associated with this mutation suggests that it functions in regulating the proliferation/differentiation of oval cells within the liver, which further indicates that this gene may function in pathological conditions that include oval cell proliferation, such as hepatocellular carcinogenesis.

Animals↗
PubMed1995

The mouse homolog of the Wiskott-Aldrich syndrome protein (WASP) gene is highly conserved and maps near the scurfy (sf) mutation on the X chromosome.

The mouse WASP gene, the homolog of the gene mutated in Wiskott-Aldrich syndrome, has been isolated and sequenced. the predicted amino acid sequence is 86% identical to the human WASP sequence. A distinct feature of the mouse gene is an expanded polymorphic GGA trinucleotide repeat that codes for polyglycine and varies from 15 to 17 triplets in different Mus musculus strains. The genomic structure of the mouse WASP gene is expressed as an approximately 2.4-kb mRNA in thymus and spleen. Chromosomal mapping in an interspecific M. Musculus/M. spretus backcross placed the Wasp locus near the centromere of the mouse X chromosome, inseparable from Gata1, Tcfe3, and scurfy (sf). This localization makes Wasp a candidate for involvement in scurfy, a T cell-mediated fatal lymphoreticular disease of mice that has previously been proposed as a mouse homolog of Wiskott-Aldrich syndrome. Northern analysis of sf tissue samples indicated the presence of WASP mRNA in liver and skin, presumably as a consequence of lymphocytic infiltration, but non abnormalities in the amount or size of mRNA present.

Amino Acid Sequence↗
PubMed1995

Ectopic expression of the agouti gene in transgenic mice causes obesity, features of type II diabetes, and yellow fur.

Mice that carry the lethal yellow (Ay) or viable yellow (Avy) mutation, two dominant mutations of the agouti (a) gene in mouse chromosome 2, exhibit a phenotype that includes yellow fur, marked obesity, a form of type II diabetes associated with insulin resistance, and an increased susceptibility to tumor development. Molecular analyses of these and several other dominant "obese yellow" a-locus mutations suggested that ectopic expression of the normal agouti protein gives rise to this complex pleiotropic phenotype. We have now tested this hypothesis directly by generating transgenic mice that ectopically express an agouti cDNA clone encoding the normal agouti protein in all tissues examined. Transgenic mice of both sexes have yellow fur, become obese, and develop hyperinsulinemia. In addition, male transgenic mice develop hyperglycemia by 12-20 weeks of age. These results demonstrate conclusively that the ectopic agouti expression is responsible for most, if not all, of the phenotypic traits of the dominant, obese yellow mutants.

Aging↗
PubMed1995

Characterization of the human homologue of the mouse Tg737 candidate polycystic kidney disease gene.

We previously identified a gene from the mutant locus in a new mouse mutation that causes recessive polycystic kidney disease. Here we describe the cloning, characterization and mapping of the homologous human gene. The human and mouse genes are 95% identical at the predicted amino acid sequence level, and both genes encode a putative protein that contains a tetratricopeptide repeat motif. The human gene, called hTg737, is expressed with a broad tissue distribution that includes the the kidney and liver, and gives rise to a 2.9 kb mRNA. The gene contains 26 exons and spans a genomic region greater than 100 kb. Chromosome mapping experiments revealed that the hTg737 gene maps near the centromere on the long arm of human chromosome 13, at position 13q12.1. While this gene does not map to the primary locus that has been identified for ARPKD in humans, it may represent a candidate gene for other recessive renal disorders that have yet to be mapped.

Amino Acid Sequence↗
PubMed1995

Early gestational expression of transforming growth factor beta isoforms by the ovine placenta.

The ruminant blastocyst metamorphoses from a 1-mm sphere to a 1 x 190-mm thread between Days 12 and 15 of gestation. The transforming growth factor beta (TGF beta) family of polypeptide growth factors are potential regulators of embryonic elongation because of their ability to regulate cellular replication, differentiation, and extracellular matrix formation. In mammalian development three isoforms of TGF beta-- TGF beta 1, beta 2, and beta 3--have been identified. Through the use of isoform-specific probes, ovine embryonic TGF beta 1 and beta 2 transcripts were identified and characterized in the present study. TGF beta 1 was expressed as a single transcript 2.6 kb in length. Levels increased by 14.8-fold from Day 13, when levels were barely detectable, to Day 27, when they were highest (p = 0.0001). Enriched samples of Day 20 chorionic and allantoic membranes demonstrated that the allantoic membrane contained 3.9-fold greater levels (p = 0.001) of TGF beta 1 message than the chorion. TGF beta 2 was expressed as five transcripts 6.2, 5.2, 4.8, 4.0, and 2.7 kb in length. From Day 13 to Day 23, there was a 6.2-fold increase (p = 0.0005) in TGF beta 2 expression; thereafter, expression declined 20% by Day 30 (p = 0.03). Extra-embryonic membrane expression of TGF beta 2 was slightly greater (1.5-fold; p = 0.01) in allantois than in chorion. Expression of TGF beta 3 was not detectable by Northern blotting, and only trace quantities of TGF beta 3 transcript were detected by slot-blot analysis. Antisera specific to TGF beta 1, beta 2 and beta 3 were used to immunolocalize the proteins within tissues. TGF beta 1 and beta 2 were identified in Day 16 trophectoderm and yolk sac. Both were also localized in chorion, allantois, and placental endothelium through Day 30. TGF beta 3 protein could not be detected in any conceptus tissue at any of the stages examined. The increase in expression of TGF beta 1 and beta 2 mRNA coincidental with conceptus elongation and placental development, as well as tissue localization of the protein, suggests their involvement in early gestational development in sheep.

Animals↗
PubMed1995

Insertional mutagenesis and molecular analysis of a new gene associated with polycystic kidney disease.

We have identified a new insertional mutation in the mouse (TgN737Rpw) that causes a phenotype that closely resembles human autosomal recessive polycystic kidney disease. The renal pathology in these mutants first presents itself as a dilation of the proximal tubules, which is quickly followed by the development of cystic lesions in the collecting ducts. The livers in the mutant animals develop a variable lesion depending upon the genetic background. We have cloned the mutant locus and have isolated and characterized a gene, Tg737, whose expression is disrupted in the mutant animals. Expression of the Tg737 gene can normally be detected using the Northern blot analysis at low levels in a variety of tissues, including the kidney and liver. Using the in situ hybridization procedure, expression of the Tg737 mRNA can be detected in the collecting ducts of adult kidneys and in portions of the embryonic day 15.5 kidney. Most important, we have corrected the defective kidney trait by expressing the wild-type cDNA as a transgene in the mutant animals. The human homologue of the Tg737 gene has also been cloned and mapped to human chromosome 13.

Animals↗
PubMed1994

CD4+CD8- T cells are the effector cells in disease pathogenesis in the scurfy (sf) mouse.

Mice hemizygous for the X-linked mutation, scurfy (sf), exhibit a fatal lymphoreticular disease that is mediated by T lymphocytes. To evaluate the respective roles of CD4 or CD8 single positive T cells in scurfy disease, neonates were treated with mAbs directed against the CD4 or CD8 molecules. Whereas mice treated with an anti-CD8 Ab developed lesions and succumbed to disease at the same time (17 days) as their untreated scurfy littermates, mice treated with an anti-CD4 Ab lived up to 11 wk before developing scurfy disease. To insure a more complete elimination of the T cell subsets, the scurfy mutation was bred onto beta 2-microglobulin (beta 2m)-deficient (CD8-less) and CD4-deficient transgenic mouse lines. Whereas there was little moderation of disease in beta 2m-deficient scurfy mice, CD4-deficient scurfy mice had markedly decreased scurfy lesions and a prolonged life span, similar to that of anti-CD4-treated sf/Y mice. Additionally, scurfy disease was transplanted into H-2-compatible nude mice through the adoptive transfer of CD4+CD8- T cells, but not CD4-CD8+ T cells. Flow-cytometric analysis revealed that sf/Y mice have an increased percentage of activated CD4+ T cells in their lymph nodes. In addition, there is an increase in the in vitro production of cytokines in the cultured splenocytes of CD8-less, but not CD4-less, scurfy mice. These data suggest that CD4+ T cells are critical mediators of disease in the scurfy mouse.

Animals↗
PubMed1994

Candidate gene associated with a mutation causing recessive polycystic kidney disease in mice.

A line of transgenic mice was generated that contains an insertional mutation causing a phenotype similar to human autosomal recessive polycystic kidney disease. Homozygotes displayed a complex phenotype that included bilateral polycystic kidneys and an unusual liver lesion. The mutant locus was cloned and characterized through use of the transgene as a molecular marker. Additionally, a candidate polycystic kidney disease (PKD) gene was identified whose structure and expression are directly associated with the mutant locus. A complementary DNA derived from this gene predicted a peptide containing a motif that was originally identified in several genes involved in cell cycle control.

Amino Acid Sequence↗
PubMed1994

Transplantation of T cell-mediated, lymphoreticular disease from the scurfy (sf) mouse.

The X-linked mutation, scurfy (sf), causes a fatal lymphoreticular disease characterized by runting, lymphadenopathy, splenomegaly, hypergammaglobulinemia, exfoliative dermatitis, Coombs'-positive anemia, and death by 24 days of age. T lymphocytes are required to mediate this syndrome as shown by a total absence of disease in mice bred to be scurfy and nude (sf/Y; nu/nu). The scurfy phenotype is not transmitted by sf/Y bone marrow transplants, though cells of scurfy origin do reconstitute all lymphoid organs in the recipient mouse. These data suggest that scurfy disease results from an abnormal T cell development process and not from an intrinsic stem cell defect. We therefore tested the ability of transplanted scurfy thymuses to transmit scurfy disease to congenic euthymic mice, to athymic (nude) mice, and to severe combined immunodeficiency (SCID) mice. Euthymic recipients of sf/Y thymic grafts remained clinically normal as did all SCID and nude recipients of normal thymus transplants. Morphological lesions similar to those found in scurfy mice occurred in all H-2-compatible nude and SCID recipients of sf/Y thymic grafts. Intraperitoneal injections of scurfy thymocytes, splenocytes, and lymph node cells also transmitted the scurfy phenotype to H-2-compatible nude mice and SCID mice. Our findings indicate that scurfy disease can be transmitted to T cell-deficient mice by engraftment of scurfy T cells, but that pathogenic scurfy T cell activities can be inhibited (or prevented) in immunocompetent recipient mice.

Animals↗
PubMed1993

Deregulation of Pax-2 expression in transgenic mice generates severe kidney abnormalities.

The Pax genes comprise a family of transcription factors active in specific tissues during embryonic development and are associated with at least three developmental mutations in mouse and man. In the developing kidney, Pax-2 is expressed in the induced mesenchyme, in the ureter epithelium, and in early epithelial structures derived from the mesenchyme. Pax-2 expression is repressed upon terminal differentiation of the renal tubule epithelium, but persists in the undifferentiated epithelium of human Wilms' tumours. We have produced a dominant gain-of-function mutation in transgenic mice by deregulating the expression of the mouse Pax-2 gene. The data obtained with four independently derived transgenic embryos and with one transgenic line demonstrate that deregulated Pax-2 expression results in histologically abnormal and dysfunctional renal epithelium with properties similar to congenital nephrotic syndrome. Thus, repression of Pax-2 is required for normal kidney development and persistent expression of Pax-2 may restrict the differentiation potential of renal epithelial cells.

Animals↗
PubMed1993

Renal failure attributable to atrophic glomerulopathy in four related rottweilers.

Atrophic glomerulopathy resulting in chronic renal failure was diagnosed in 4 related Rottweilers, each < 1 year old. All 4 dogs had severe azotemia and massive protein-losing nephropathy. Histologically, the glomerular lesion was characterized by mild dilatation of Bowman's space, with glomerular tufts absent or markedly atrophied. The lesion is distinct from the congenital glomerular changes described in Samoyeds or Doberman Pinschers.

Animals↗