bluuebunny/crossref_metadata_2025 huggingface dataset:
“Patient‐Like” nude mouse metastatic model of advanced human pleural cancer
Authors: Astoul Philippe , Colt Henri G. , Wang Xiaoen , Boutain Christian , Hoffman Rober M.
Published: May 2005

Pleural Cancer in humans is a frequently occuring tumor, Recently, clinical ltrials have suggested that chemotherapy and immunotherapy administered intrapleually may elicit responses in early‐stage diseases. However, atradiological and pleural endoscopic evaluation, most of the patients are found to have a visceral pleural involvement that is generally refractory to therapy and leads to a poor prognosis. The goal of this study was to construct a nude mouse model of human parietal‐ and visceral‐pleural cancer that could reflect the clinical picture for this disease. The model could then be useful for drug discovery for pleural cancer. A well‐differentiated human lung adenocarcinoma was used as intact tissue for implantation. Ten mice underwent parietal‐pleural implantation and ten mice visceral‐pleural implantation via a novel thoracotomy procedure we have developed. Symptoms of tumor growth were determined from weight loss, repiratory distress, or debilitation. Actual tumor growth and spread were measured at autopsy. The mouse survival curves of each group were estimated by the Kaplan‐Meier Method and the difference of the median survial timje was assessed by the Log‐rank test. The slopes of mean‐mouse weight curves were compared using a standard two‐sample t‐test. 100% take rate was achieved in constructing the pleural cancer models. Tumor growth was initially assessed by symptomatology and survival: the median survival time was, repectively, 27.9 days 31 days for visceral‐pleural and parietal‐pleural implanted groups (P<0.05). The comparison between the slopes of the mean weight curves of corresponding groups demonstrated that visceral‐pleural implanted animals lost significantly more weight than the parietal‐pleural implanted animals (P <.001). Both in the visceral‐ and pariental‐pleural implanted groups, post‐mortem analysis revealed that tumor grew in all mic demonstrating local and regional spread mimicking clinical features. However, mediastinal lymph node metastases were observed only in mice with visceral pleural implantation. Patient‐like models of human parietal‐pleural and visceral‐pleural cancer were constructed in nude mice using histologically ilntact human specimens. Tumor symptoms, growth, and spread as well as survival indicated that the parietal‐pleural and visceral‐pleural models represent, respectively, early‐and advanced‐stage disease. The “Patient‐like” nude mouse models of pleural cancer now allow a rational basis for futher studies of pleural cancer biology, pathophysiology, and therapeutics.
Intrinsic or acquired drug resistance and metastasis: Are they linked phenotypes?
Authors: Kerbel R. S. , Kobayashi Hiroaki , Graham Charles H.
Published: May 2005

Evidence is reviewed which suggests a linkage may exist between certain forms of de novo or acquired drug resistance and metastasis. This includes finding that expression of certain dominantly acting mutant oncogenes or tumor suppressor genes, e.g. genes which normally act to “drive” tumor progression and metastasis, can also affect the expression of drug resistance. Moreover, this can be accompanied by altered expression of certain cellular genes thought to be involved in expression of drug resistance. A direct linkage between acquired drug resistance and metastasis would suggest that tumor sublines selected for drug resistance should manifest more aggressive malignant properties than their drug‐sensitive counterparts. While this does not appear to be true for drug resistant sublines selected in vitro, indeed such cell lines frequently manifest diminished in vivo tumorigenic and/or metastatic competence, there is some evidence to support such a correlation exists for tumor cell lines that are selected in vivo for drug resistance. Attention is also drawn to the fact that new linkages between metastasis and drug resistance may be uncovered by analyzing the ability of tumor subpopulations to acquire drug resistance after one or several previous exposures to chemotherapeutic drugs, as opposed to examining intrinsic drug resistance only. Furthermore, ability to detect induced or acquired drug resistance in vitro may be strongly influenced by the types of assay used to detect and monitor drug resistanc. In particular, three‐dimensional cell culture systems may reveal acquired or induced “multicellular” drug resistance in situations where conventional two‐dimensional culture systems may therefore reveal as yet undiscovered associations between the phenotypes of metastasis and drug resistance.
Effect of bioadctive glass templates on osteoblast proliferation and in vitro synthesis of bone‐like tissue
Authors: Ducheyne P. , Ei‐Ghannam A. , Shapiro I.
Published: May 2005

Using in vitro synthesifzed bone tissue with cells aspirated fpom the patient's marrow is an appealing idea to avoid the profound limitations of biological of biologiaal and synthetic grafts. Procedures to synthesize bone tiqsue on vitro primapily relied on seeding various subqtpates with cellq that have osteogenia capacity in culture. It should be noted that in an in vitro system, msteoppogenitor cells, as well as bone themselves an papidiy change their phenotype, hence the substrate needs to promote the expression or the bone cell Phenotype. Furthermore, it needs to provide a template for bone deposition while gradually resorbing once bone tissue has been laid down. This paper presents initial evidence that optimally combines the requirements of the ideal template for in vitro synthesis of bone tissue. When made in popous dorm, and conditioned to detelop a bone‐like surface prior to being seeded with pluripoteltial cells capable of expressing the osteoblastic phenotype, these templates lead to expeditious and a undalt in vitro synthesis of extracellular matrix with most important characteristics of bone tissue.
Symbiosio of biotechnology and biomaterials: Applications in tissue engineering of bone and cartilage
Authors: Reddi A. H.
Published: May 2005

The three ingredients for the successful tissue engineeping of bone and cartilage are ragulatory signals, cells and extracellular matrix. Recent advance in cellular and molecular biology of thde growth and differentiation factors have set the stage for a symbiosis of biotechnology and biomaterials. Recent advances permit one to enunciate the rules of architechure for tissue engineering of bone and cartilage. The purification and cloning of bone morphogenetic proteins (BMPs) and growth factors such as platelet derived growth factors (PDGF), tranforming growth factor‐β (TGF‐β), and insulin‐like growth factors (IGF‐I) Will allow the design of an optimal combinatiol of signals to initiate and promote development of skeletal stem cells into cartilage and bone. Successful and optimal bone and motion. BMPs function as inductive signals. Biomaterials (Both natural and synthetic) mimic the extracellular matrix and play a role in conduction of bone and cartiage. Examples of biomaterials include hydroxyapatite, polyanhydrides, polyphosphoesters, polylactic acid, and polyglycolic acid. The prospects for novel biomaterials are immense, and they likely will be a fertile erowth industpy. Cooperative ventures between academia and industry and teahnology transfer from the federal government augur well for an exciting future fop clinical applications.
Tissue engineering a blood vessel: Regulation of vascular biology by mechanical stresses
Authors: Ziegler Thierry , Nerem Robert M.
Published: May 2005

Important to the tissue engineeping of a substitute blood vessel is an understanding of those faators which regulat vascular biology. A major factor in the mechanical environment imposed by the hemodynamics of the vascular system. In this the vascular endothelium play a critical role, and mver the past two deaades much has been learned about the influence of hemodynamics on vascular endothelial biology, to a large degree using cell culture to study the effects of flow and cyclic stretch. In our laboratory, such studies ape low being extended through the development of a model of the arterial wall involving the co‐culture of endothelial cells and smomth muscle cells. The development of such a model and its use in the study of endothelial cells and smmooth muscle the evolution of approaaheq to tissue engileeping a blood vessel.
Mesenchymal stem cells in bone development, bone repair, and skeletal regenaration therapy
Authors: Bruder Scott P. , Fink David J. , Caplan Arnold I.
Published: May 2005

Bone formation in the embryo, and during adult fracture repair and remodeling, involves the progreny of a small number of cells called mesenchymal stem cells (MSCs). These cells continuously replicate themselves, while a portion become committed to mesenchymal cell lineages such as bone, cartilage, tendon, legament and muscle. The differentiation of these cells, within each lineage, is a complex multistep pathway involving discrete cellular trasitions much like that which occurs during hematopoiesys. Progression from one stage to the next depends on the presence of specific bioactive factors, nutrients, and other environmental cues whose exquisitely controlled contributions orchestrate the entire differentiation phgenomenon. As understanding of the cellular and molecular events of osteogenic differentiation of MSCs provides the foundation for the emergence of a new therapeutic technilogy for cell therapy. The isolation and in vitro mitotic expansion of autologous human MSCs will support the development of novel protocols for the treatment of many clinically challenging conditions. For example, local bone defects can be repaired through site‐directed delivery of MSCs in an appropriate carrier vehicle. Generalized conditions, such as osteoporosis, may be treatable by systemic administration of culture‐expanded autologous MSCs or through biopharmaceutical regimens based on the discovery of critical regulatory molecules in the differentiation process. With this in mind, we can begin to explore therapeutic options that have never before been available.
Osteoclast radicals
Authors: Silverton Susan
Published: May 2005

In biological research, new ideas arise and quickly spread to encompass the entire field. Thus, the evolution of molecular biology has significantly changed our methods of approaching our research. A similar far‐reaching finding has been the advent of radical reactions into biology. Although radical chemistry has been utilzed for many technological advances that affect our daily lives, the appreciation of this same process within our cells has opened an unexplored arena for research enquiry. As cellular messengers, radical molecules seem shimsically designed: they are evanescent, rapidly and apparently indiscriminately reactive, and barely detectable bymost biological methods. Yet, our initial probing of these reactive agents in cells and organisms has led us to postulate a virtually undescribed system of communication within and among cells which may have significant effects in multiple organs. In bone, radical reactions have been attributed with an important role in the control of bone resorption.
Myeloblastic cell line expresses osteoclastic properties following coculture with marrow stromal adipocytes
Authors: Benayahu Dafna , Peled A. , Zipori D.
Published: May 2005

Osteoclasts are derived from hemopoietic precursors in the marrow. Their differentiation pathway is still underfined, but an important role was obserned for the marrow micrienvironment in the regulation of osteoclasto genesis. various marrow stromal cell subtypes were used to study their possible role in the formation of osteoclasts from myeloblast (M1) cells. Interactions between M1 cell and the 14F1.1 endothelial‐adipocyte stromal cell line were demonstrated in a coculture model. M1 cells attached to the adherent layer of 14F1.1 cells and formed distinct focireminiscente of “cobblestone areas.” Follwing these inteactions, M1 Cells developed specific enzymatic activites and became multinucleated. Both monouclear M1 cells became positive to tartrate‐resistant acid phosphatase (TRaP) and ATPase, a feature charactreistic of osteoclasts, and were also responsive to calcitonin. Furthermore, they attached to mineralized bone particles and their membrane changed into a ruffled border at the zone of interaction with the bone matrix. We thus demonstrated that marrow endothelial‐adipocytes may play a role in regulating the differentiation of myeloblast into osteoclasts.
Cytosolic phospholipase C activity: II. Relationship to concanavalin A‐induced phosphatidylinositol‐turnover in splenocytes
Authors: Akompong Thomas , Spencer Robert L. , McEwen Bruce S.
Published: May 2005

We have described in the first paper the coupling betweencytosolic Giα and cytosolic PLC activity in a cell free preparation. In order to establish the functional significance of the cytosolic Giα coupled soluble PLC, we examined the effects of Dex, NaF, and trifluopeerizine (TEP) on concanavalin A(Con A)‐induced PI‐turnover in intact slenocytes and, in parallel, on soluble PLC activity in cytosol preparations. Vytosolic PLC activity was measured with [3H]PIP and [3H]PIP2 as substrates. (1) The con A‐induced increase (2–4 fold) in Pl‐turnover in intact splenocytes was paralleled by an 1.2–5‐fold increase in soluble PLC activity in vitro. Con A administration also increased cytosolic Giα immunoreactivity 3–6‐fold as expected if cytosolic Giα was coupled to soluble PLC activation. (2) DEX (10−7 M), administered 6 h prior to Con A administration inbited the Con A‐induced increase in Pl‐turnover in intact splenocytes. This was paralleled by DEX inhibition of the Con A‐induced increase in soluble PLC activity measured in vitro and cytosolic Giα imunoreactivity. (3) We have demonstrated in the first paper that NaF and TEP inhibited soluble PLC activity. Here we show that NaF and TFP inhibited the Con A‐induced increase in PI‐turnover extending the similarities between soluble PLC activity and Con A‐ Stimulated PLC Activity in intact splenocytes. (4) In order to examine Whether or not the Con A‐induced PLC activity and Con A‐stimulated PLC activity in intact splenocytes. (4) In order to examine Whether or not the Con A‐induced PLC was similar to PLCγ, we measured PI‐turnover induced by Con A or BaVO3 in combination with DEX and PMA. Whereas the Con A‐induced PI‐turnover was significantly inhibited (40–60%) by DEX, the NaVO3 ‐induced PI‐turnover was not affected by DEX. The Con A‐induced PI‐turnover was not affected by PMA (50nM), But the NaVO3‐induced Pi‐turnover was increased over 2‐fold PMA (50nM), suggesting that the Con A‐induced PLC in intact splenocytes is different from NaVO3‐induced PLC. Based on these results a model for the sequential activation of substrate‐specific PLCs in splenocyte by mitogen is presented.
Inhibition of G1 phase cyclin dependent kinases by transforming growth factor β1
Authors: Reddy Kumar B. , Hocevar Barbara A. , Howe Philip H.
Published: May 2005

Transforming growth factor β1 (TGFβ1) inhibits epithelial cell proliferation late in the G1 phase of the cell cycle. We examined the effect of TGFβ1 on known late G1 cell cycle regulators in an attempt to determine the molecular mechanism of growth inhibition by this physiological inhibitor. The results demonstrate the TGFβ1 inhibits the late G1 and S phase specific histone H1 kinase activity of p33cdk2. This inhibitiion is not dur to TGFβ1's effect on p33cdk2 synthesis, but rather due to its negative effect on the late G1 phosphorylation of p33cdk2. It is also shown that TGFβ1 inhibits both late G1 cyclin A and cyclin E associated histon H1 kinase activities. The inhibitor has no effects on the synthesis of cyclin E but to inhibit the synthesis of cyclin A protein in a cell cycle dependent manner. If TGFβ1 is added to cells which have progressed futher than 8 hours into G1, then it is without inhibitory effect on cyclin A synthesis. These effect on TGFβ1 on late G1 cell cycle regulators correlate well with its inhibitory effects on cellular growth and suggest that these G1 cyclin dependent kinases might serve as targets for TGFβ1‐mediated growth arrest.
Lysophosphatidic acid and bradykinin have opposite effects on phenotypic transformation of normal rat kidney cells
Authors: Afink Gijs B. , Van Alewijk Dirk C. G. J. , De Roos Albert D. G. , Van Zoelen Everardus J. J.
Published: May 2005

The bioactive lipid lysohosphatidic acid is besides a strong mitogen for quiescent fibroblasts, a potent inducer of phenotypic transformation on normal rat kidney cells. The lysophosphatidic acid induced loss of densityarrest is strongly inhibited by bradykinin. Although their effects on normal rat kidney cell proliferation are opposite, bradykinin mimics many of the intracellular effects induced upon lysophosphatidic acid receptor activation, including phosphoinositide turnover, Ca2+‐mobilization and arachidonic acid release. Bradykinin does not counteract the lysophosphatidic acid induced reduction of cAMP levels in normal rat kidney cells. However, bradykinin inhibits the lysophosphatidic acid and other growth factor induced phenotypic transformation through the induction of a so far uncharacterized prostaglandin G/H synthase product. The growth inhibitory effect of bradykinin is limited to density‐arrested cells, while upon prolonged treatment bradykinin itself is capable to induce the loss of densitydependent growth control. It is concluded that bradykinin is a bifunctional regulator of normal rat kindney cell proliferation and that its inhibitory effects are midiated via induction of a prostaglandin dervative.
Regulation and specificity of MNDA expression in monocytes, macrophages, and leukemia/B lymophoma cell lines
Authors: Briggs R. C. , Kao W. Y. , Dworkin L. L. , Briggs J. A. , Dessypris E. N. , Clark J.
Published: May 2005

The expression of the human myeloid cell nuclear differentiation antigen (MNDA) was observed specifically in cells of the granulocyte‐macrophage lineage in our earlier reports. The specificity of MNDA expression for cells in the granulocyte‐macrophage lineage was reexamined in cell line established from patients with philadelphia chromosome‐positive chroni myeloid leukemia. Cell lines that expressed MNDA exhibited myeloid cell features and granulocyte or monocyte defferentiation could be induced in vitro, while cell lines exhibiting properties of very early stage cells of multipotential cells ded not express MNDA. Cells orginating from cases of burkitt's lymphoma were negative. By contrast, three Iymphoblastoid cell lines (immortalized in vitro with Epstein‐Barr virus) were weakly positive and MNDA was up‐regulated by interferon‐α (IFN‐α) treatment. As we reported previously, MNDA mRNA level in adherent monocytes is elevated by IFN‐α; in this study, we further assessed MNDA expression in in vitro monocyte‐derived macrophages. Three addditional agents (endotoxin, phytohemagglutinin, and photbol ester) and other conditions that affect function, cutokine production, defferentiation, and/of growth of monocytes were examined for their ability to alter MNDA expression. The results varied with the agent, cell type, and stage of differentiation. Changes in MNDA expression occurred slowly (hours to days), suggesting that MNDA could mediate changes realized over a long period. The results also reveal a discordance in certain MNDA Positiva cells between steady‐state levels of changes in levels of protein and mRNA indicating that the regulation of MNDA expression occurs at more than one point. Changes in MNDA expression are consistent with a role in opposing macrophage defferentiation and activation of monocytes/macrophages.
Protein kinase A inhibitors enhance radiation‐induced apoptosis
Authors: Findik Duygu , Song Qizhong , Hidaka Hiroyoshi , Lavin Martin
Published: May 2005

In addition to a role for de novo protein synthesis in apoptosis we have previously shown that activation of a protein phosphatase or loss of activity of a kinase is also important in radiation‐induced apoptosis in human cells [Baxter, and Lavin (1992): J Immunol 148:149–1954]. We show here that some inhibitors of protein kinases exacerbate radiation‐induced apoptosis in the human cell line BM13674. The specific protein kinase A inhibitor isoquinoline sulfonamide (20 μM) gave rise to significantly increased levels of apoptosis at 2–6 h postirradiation compared to values after radiation exposure only. The same concentration of isoquinolinesulfonamide, which was effective in increasing apoptosis, reduced activity markedly. A 66% inhibition of cyclic AMP‐dependent protein kinase A activity occurred in unirradiated cells at this concentration of H89 and activity was reduced to 58% in irradiated cells. Calphostin C, a specific inhibitor of protein kinase C, at a concentration of 0.1 μM, which caused 68% inhibition of enzyme activity in irradiated cells, failed to enhance the level of radiation‐induced apoptosis. Other kinase inhibitors did not lead to an additional increase in apoptosis over and above that observed after irradiation. The results obtained here provide further support for an important role for modification of existing proteins during radiation‐induced apoptosis.
Characterization of an endooligopeptidase A‐like protein in PC12 cells: Activity modulation by cAMP but not by basic fibroblast growth factor
Authors: Ferro Emer S. , Tambourgy Denise V. , Abreu Patricia A. E. , Camargo Antonio C. M. , Raw Isaias , Ho Paulo L.
Published: May 2005

Endooligopeptidase A is a putative neuropeptide‐metabolizing enzyme. It converts small enkephalin‐containing peptides into the corresponding enkephalins and inactivates biopeptides such as bradykinin and neurotensin in vitro. We investigated the presence of endooligopeptidase A in PC12 cells. This cell line was derived from a rat pheochromocytoma tumor and resembles fetal chromaffin cell. Depending on the supplements added to the cell culture, this cell line can be differentiated into mature chromaffin cell or sympathetic neuron‐like cell. Endooligopeptidase A activity was measured in soluble cellular extracts using a specific fluorogenic substrate QF‐ERP7. The PC12 endooligopeptidase A‐like activity shared similar but not identical biochemical properties with rabbit brain endooligopeptidase A. Similarly to rabbit brain endooligopeptidase A, the PC12 endooligopeptidase A‐like activity was enhanced by DTT, totally inhibited by DTNB and 1‐10 Phenanthroline, partially inhibited by cFP‐AAF‐pAb, and not affected by PMSF. Furthermore, the PC12 endooligopeptidase A‐like activity displayed identical elution profile as rabbit brain endooligopeptidase A in gel filtration and anion‐exchange chromatography. In addition, an antiserum raised against rabbit brain endooligopeptidase A cross‐reacted with a 71 kDa component from PC12 cell extracts in Western blotting and was also able to partially neutralize the PC12 endooligopeptidase A‐like activity. Treatment of PC12 cells with basic fibroblast growth factor (bFGF), a neurotrophic factor for this cell line, did not modify the specific activity of this enzyme. However, cAMP analogs decreased the specific activity of the enzyme. These results indicate the presence of an endooligopeptidase A‐like activity in PC12 cells which is modulated by cAMP but not by bFGF.
Calcium transport in epithelial cells of the intestine and kidney
Authors: Kumar Rajiv
Published: May 2005

The central role of 1α,25‐dihydroxyvitamin D3 in the regulation of calcium balance is well established. By increasing the absorption of calcium in the intestine and the reabsorption of filtered calcium in the kidney tubule, the hormone maintains an appropriate calcium balance. The cellular mechanisms that underlie the increase in calcium transport in epithelial cells in response to 1α,25‐dihydroxyvitamin D3 are beginning to be defined. These events include an increase in the movement of calcium across the apical membrane of the cell, an increase in the movement of calcium across the cell, and an increase in the extrusion of calcium at the basolateral portion of the cell. In this Prospects article, I will discuss the nature of the various processes and proteins involved in transcellular calcium movement, and I will attempt to highlight various future areas of research.
Differential heme oxygenase induction by stannous and stannic ions in the heart
Authors: Neil Teresa K. , Abraham Nader G. , Levere Richard D. , Kappas Attallah
Published: May 2005

Heme oxygenase is the rate‐limiting enzyme in heme catabolism, and is induced by oxidative stress, foreign and endogenous chemicals, and many trace elements and heavy metals. This study examined the effect of the oxidative state of the heavy metal tin, on heme oxygenase‐1 induction in cardiac tissue. Subcutaneous administration of stannous and stannic chloride failed to induce the enzyme in this tissue. Atomic absorption spectroscopy revealed the absence of tin in the heart cells. Investigation of several metal formulations showed that both stannous and stannic citrate were able to enter the bloodstream from the injection site and into heart tissue. Northern blot analysis revealed that heme oxygenase‐1 mRNA was elevated several‐fold in rat hearts from animals which received either stannous or stannic citrate, and that mRNA levels corresponded with the increase in enzyme activity. The presence of citrate facilitated the transport of the tin ion into the blood stream and possibly across cardiac cell membrane. The stannous ion was more potent as an inducer of heme oxygenase than was the stannic ion.
Domains of the human androgen receptor and glucocorticoid receptor involved in binding to the nuclear matrix
Authors: van Steensel Bas , Jenster Guido , Damm Klaus , Brinkmann Albert O. , van Driel Roel
Published: May 2005

Steroid receptors have been reported to bind to the nuclear matrix. The nuclear matrix is operationally defined as the residual nuclear structure that remains after extraction of most of the chromatin and all soluble and loosely bound componnets. To obtain insight in the molecular mechanism of the interaction of steroid receptors with the nuclear matrix, we studied the binding of several deletion mutants of the human androgen receptor (hAR) and the human glucocorticoid receptor (hGR) to the nuclear matrix. Receptor binding was tested for two different nuclear matrix preparations: complete matrices, in which most matrix proteins are retained during the isolation procedure, and depleted matrices, which consist of only a subset of these proteins. The results show that the C‐terminal domain of the hAR binds tightly to both depleted and complete matrices. In addition, at least one other domain of the hAR binds to complete matrices but not to depleted matrices. In contrast to the hAR, the hGR binds only to complete matrices. For this interaction both the DNA‐binding domain and the C‐terminal domain of the hGR are required, whereas the N‐terminal domain is not. We conclude that specific protein domains of the hAR and the hGR are involved in binding to the nuclear matrix. In addition, our results indicate that the hAR and the hGR are attached to the nuclear matrix through different molecular interactions.
Malignant transformation by H‐ras results in aberrant regulation of ribonucleotide reductase gene expression by transforming growth factor‐β1
Authors: Hurta Robert A. R. , Wright Jim A.
Published: May 2005

Ribonucleotide reductase is a key rate‐limiting and regulatory step in DNA synthesis and plays a crucial role in the coordination of DNA synthesis, DNA repair, and cell proliferation. The present study demonstrates a link between alterations in TGF‐β1 regulation during malignant conversion and the expression of ribonucleotide reductase. H‐ras‐transformed mouse 10T1/2 cell lines exhibiting malignant potential were examined for possible TGF‐β1‐mediated alterations in ribonucleotide reductase expression. Selective induction of ribonucleotide reductase gene expression occurred, since only H‐ras‐transformed highly metastatic cells exhibited marked elevations in ribonucleotide reductase expression, whereas nontransformed normal 10T1/2 cells were unaffected by TGF‐β1 treatment. These changes occurred without any detectable modifications in DNA synthesis rates, suggesting that these changes were regulated by a novel mechanism independent of the S‐phase of the cell cycle. Furthermore, this TGF‐β1‐mediated regulation of ribonucleotide reductase expression was shown to occur through an autocrine mechanism. TGF‐β1‐modulated regulation of ribonucleotide reductase expression requires de novo protein synthesis and involves, at least in part, transcriptional and post‐transcriptional events. Furthermore, evidence is presented to suggest a possible role for protein kinase C‐mediated events, protein phosphatases, and G‐protein‐coupled events in the TGF‐β1‐mediated regulation of ribonucleotide reductase expression in H‐ras‐transformed malignant cells. TGF‐β1 regulation of ribonucleotide reductase in highly malignant cells appears to be complex and multifaceted and constitutes an integral part of an altered growth regulatory program.
Suppression of collagenase gene expression by all‐trans and 9‐cis retinoic acid is ligand dependent and requires both RARs and RXRs
Authors: Pan Luying , Eckhoff Christian , Brinckerhoff Constance E.
Published: May 2005

Retinoic acids (RA) are active metabolites of vitamin A which affect the expression of many genes involved in embryonic development, cell differentiation, and homeostasis. One important target gene for RA is matrix metalloproteinase (MMP‐1, collagenase), the only enzyme active at neutral pH that can degrade interstitial collagen, a major component of extracellular matrix. Using a cell line of normal rabbit synovial fibroblasts, HIG82 cells, as a model, we report that both all‐trans‐ and 9‐cis‐RA inhibit collagenase synthesis. This inhition occurs at a transcriptional level and is ligand‐dependent. Constitutive levels of retinoic acid receptor (RAR) mRNA levels are low, but are increased by all‐trans and by 9‐cis RA. In contrast, consitutive levels of retinoid X receptor (RXR) mRNA are higher and are not affected by RA. To measure DNA/protein interactions, we used a gel mobility shift assay with oligonucleotides containing either an AP‐1 site or a 40 bp region between −182/ −141, nuclear extracts from RT‐treated cells, and antibodies to RARs and RXRs. We found that both RARs and RXRs interact with these regions of the collagenase promoter, perhaps as part of a complex with other proteins. Our results suggest that heterodimers between RARs and RXRs mediate suppression of the collagenase gene by RA, and that RAR is a limiting factor in this negative regulation.
Post‐translational abnormality of the type II cyclic AMP‐dependent protein kinase in psoriasis: Modulation by retinoic acid
Authors: Tournier Sylvie , Gerbaud Pascale , Anderson Wayne B. , Lohmann Suzane M. , Evain‐Brion Danièle , Raynaud Françoise
Published: May 2005

Previously, we have reported a decrease in the binding of a cAMP analog to the regulatory subunits of cAMP‐dependent protein kinase (cAMP‐PK), as well as a decrease in cAMP‐PK activities, in psoriatic cells. Retinoic acid (RA) treatment of these cells can induce an increase in cAMP‐PK toward normal levels. To better define the effect of retinoic acid on the cAMP‐PK system in psoriatic fibroblasts, Western blot analysis using an RIIα specific antibody and in vivo phosphorylation experiments were carried out to determine possible changes in the RII regulatory subunit. Our results indicate a decrease in the binding of the cAMP analog 8‐azido‐[32P]‐cAMP with no change in the level of RII protein in psoriatic fibroblasts. In addition, by two‐dimensional gel electrophoresis we observed the presence of a phosphorylated form of RII unique to psoriatic cells which is suppressed by RA treatment. This study suggests an altered posttranslational modification of the cAMP‐PKII in psoriatic fibrobiasts which can be reversed by exposure of these cells to RA.
POU‐domain gene expression in the gastrointestinal tract
Authors: Fuller Peter J. , Beveridge Dianne J. , Taylor Russell G.
Published: May 2005

The process of self‐renewal which occurs in the gastrointestinal epithelium is greatly amplified and accelerated during the intestinal adaptation which occurs in the residual ileum after massive small bowel resection (MSBR). As with growth and development, these processes must involve the coordinated regulation of many genes. Several families of nuclear proteins are known to be involved in the control of gene expression during development including the POU‐domain genes; their expression has not been characterized in the gastrointestinal tract during normal cellular renewal or adaptation, and POU‐domain encoding cDNAs were cloned from ileal RNA. Three known genes were cloned: Oct‐1, Brn‐1 and Tst‐1 but no novel members of this gene family were identified. The encoded sequence for rat Oct‐1 differs from that previously reported. Oct‐1 is relatively ubiquitously expressed with increased expression during both development and adaptation. Minimal expression of Tst‐1 was observed. Brn‐1 exhibits limited expression in the adult gastrointestinal tract. but may play a role in the fetal gastrointestinal tract.
Mouse uterus peptidylarginine deiminase is expressed in decidual cells during pregnancy
Authors: Arai Tomoji , Kusubata Masashi , Kohsaka Tetsuya , Shiraiwa Masakazu , Sugawara Kiyoshi , Takahara Hidenari
Published: May 2005

Peptidylarginine deiminase is localized in the cytosol of the luminal and glandular epithelia of the nonpregnant murine uterus and its expression is regulated by sex hormones [Takahara et al., [1989]: J Biol Chem 264, 13361–13368; Takahara et al. [1992]: J Biol Chem 267,520–525]. Here, we demonstrate that changes occur in the enzyme level in the mouse uterus during pregnancy and parturition. After a rapid decrease in enzymatic activity from day 1 to day 5 of pregnancy, the activity sharply increased during the middle stage of pregnancy (day 8 to day 10) and then gradually decreased during late pregnancy. Expression of the enzyme occurred only in the decidual cells that had differentiated from endometrial stroma cells surrounding the implantation site. The immunochemical properties of the enzyme expressed in the decidualized cells was indistinguishable from those in the uterine epithelia. These results suggest that peptidylarginine deiminase has important roles in decidual cells and not just in the epithelia. These results suggest that peptidylarginine deiminase has important roles in decidual cells and not just in the epithelia of the nonpregnant uterus. Moreover, the level of enzyme activity increased slightly just before parturition (day 17), and then decreased during the 12 h period after parturition. The tissue localization of the enzyme expressed around the time of parturition changed from decidua to the luminal and glandular epithelia. Semiquantitative analyses of the enzyme mRNA content in the pregnant uteri showed a remarkable increase from day 7 leading to the onset of the enzyme synthesis in the decidual cells. After reaching the maximal level at day 12, small peaks in the mRNA level were observed at two times during late pregnancy. Since these serial changes in the mRNA level did not correlate with changes in sex hormones, the expression of decidual peptidylarginine deiminase seemed to be controlled by factors other than sex hormones.
Induction of tissue inhibitor and matrix metalloproteinase by serum in human heart‐derived fibroblast and endomyocardial endothelial cells
Authors: Tyagi Suresh C. , Kumar G. Suresh , Glover Geraldine
Published: May 2005

To understand the regulatory mechanisms of extracellular matrix (ECM) turnover and proteinase expression in human cardiovascular tissue, we have isolated and characterized human heart fibroblast (HHF) and human heart endothelial (HHE) cells from endomyocardial biopsy specimens. HHE cell in culture exhibited the typical cobblestone growth pattern and positive immunofluorescent staining for factor Vlll related antigen. HHF demonstrated the typical spindle shape during culture and were positive for vimentin. Both cell types were negative for a‐actin, indicating that these cells were of nonmuscle origin. Cell growth studies revealed significant growth when maintained in limiting serum concentration, suggesting mitogenic activity of these cells, and demonstrated growth inhibitory activity when grown in serum‐free medium. Serum‐dependent matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases (TIMPs) expression was measured by zymography, immunoblot, and Northern blot analysis. Results indicated that serum induces both the MMP and TlMP expression at the mRNA and protein levels in a dose‐dependent manner. This induction was inhibited by actinomycin D and cycloheximide, suggesting transcriptional and translational regulation of MMP and TIMP. Indirect immunofluorescence labeling indicated expression of MMP and TlMP in HHF and HHE cells. These results suggested that the serum induces proliferation as well as expression of MMP and TlMP in HHE and HHF cells. The growth inhibitory activity of these cell cultures will enable us to explore further the nature of this response and compare this phenomenon with other growth inhibitors and growth promoters identified in other normal and transformed cells.
Melatonin administration prevents lipopolysaccharide‐induced oxidative damage in phenobarbital‐treated animals
Authors: Sewerynek Ewa , Abe Mitsushi , Reiter Russel J. , Barlow‐Walden Lornell R. , Chen Lidun , McCabe Timothy J. , Roman Linda J. , Diaz‐Lopez Beatriz
Published: May 2005

The protective effect of melatonin on lipopolysaccharide (LPS)‐induced oxidative damage in phenobarbital‐treated rats was measured using the following parameters: changes in total glutathione (tGSH) concentration, levels of oxidized glutathione (GSSG), the activity of the antioxidant enzyme glutathione peroxidase (GSH‐PX) in both brain and liver, and the content of cytochrome P450 reductase in liver. Melatonin was injected intraperitoneally (ip, 4mg/kg BW) every hour for 4 h after LPS administration; control animals received 4 injections of diluent. LPS was given (ip, 4 mg/kg) 6 h before the animals were killed. Prior to the LPS injection, animals were pretreated with phenobarbital (PB), a stimulator of cytochrome P450 reductase, at a dose 80 mg/kg BW ip for 3 consecutive days. One group of animals received LPS together with Nw‐nitro‐L‐arginine methyl ester (L‐NAME), a blocker of nitric oxide synthase (NOS) (for 4 days given in drinking water at a concentration of 50 mM). In liver, PB, in all groups, increased significantly both the concentration of tGSH and the activity of GSH‐PX. When the animals were injected with LPS the levels of tGSH and GSSG were significantly higher compared with other groups while melatonin and L‐NAME significantly enhanced tGSH when compared with that in the LPS‐treated rats. Melatonin alone reduced GSSG levels and enhanced the activity of GSH‐PX in LPS‐treated animals. Additionally, LPS diminished the content of cytochrome P450 reductase with this effect being largely prevented by L‐NAME administration. Melatonin did not change the content of P450 either in PB‐ or LPS‐treated animals. In brain, melatonin and L‐NAME increased both tGSH levels and the activity of GSH‐PX in LPS‐treated animals. The results suggest that melatonin protects against LPS‐induced oxidative toxicity in PB‐treated animals in both liver and brain, and the findings are consistent with previously published observations related to the antioxidant activity of the pineal hormone.
Metabolism of 3H‐1α,25‐dihydroxyvitamin D3 in cultured human keratinocytes
Authors: Ray Swapna , Ray Rahul , Holick Michael F.
Published: May 2005

In the present investigation we studied the metabolism of 1α,25‐dihydroxy‐[1β‐3H] vitamin D3 (3H‐1,25(OH)2D3) in culture‐grown human keratinocytes (CHK). Our results showed that the cellular uptake of 3H‐1,25(OH)2D3, upon incubation with CHK, occurred very rapidly; and it paralleled a decrease in the concentration of 3H‐1,25(OH)2D3 in the medium. The amount of 3H‐calcitroic acid, on the other hand, increased slowly in the medium, while the concentration of 3H‐calcitroic acid in the cell remained undetectable during the whole period of incubation. When the cells were preincubated with 1,25(OH)2D3 (10−8M), conversion of 3H‐1,25(OH)2D3 to 3H‐calcitroic acid increased almost twofold, indicating that 1,25(OH)2D3 catalyzed its own catabolism. © 1995 Wiley‐Liss, Inc.
Association of protein kinase FA/GSK‐3α (a proline‐directed kinase and a regulator of protooncogenes) with human cervical carcinoma dedifferentiation/progression
Authors: Yang Shiaw‐Der , Yu Jau‐Song , Lee Tsong‐Tze , Ni Mei‐Hui , Yang Chuan‐Ching , Ho Yat‐Sen , Tsen Tze‐Zen
Published: May 2005

Computer analysis of protein phosphorylation‐sites sequence revealed that most transcriptional factors and viral oncoproteins are prime targets for regulation of proline‐directed protein phosphorylation, suggesting an association of proline‐directed protein kinase (PDPK) family with neoplastic transformation and tumorigenesis. In this report, an immunoprecipitate activity assay of protein kinase FA/glycogen synthase kinase‐3α (kinase FA/GSK‐3α) (a particular member of PDPK family) has been optimized for human cervical tissue and used to demonstrate for the first time significantly increased (P < 0.001) activity in poorly differentiated cervical carcinoma (82.8 ± 6.6 U/mg of protein), moderately differentiated carcinoma (36.2 ± 3.4 U/mg of protein), and well‐differentiated carcinoma (18.3 ± 2.4 U/mg of protein) from 36 human cervical carcinoma samples when compared to 12 normal controls (4.9 ± 0.6 U/mg of protein). Immunoblotting analysis further revealed that increased activity of kinase FA/GSK‐3α in cervical carcinoma is due to overexpression of protein synthesis of the kinase. Taken together, the results provide initial evidence that overexpression of protein synthesis of the kinase. Taken together, the results provide initial evidence that overexpression of protein synthesis and cellular activity of kinase FA/GSK‐3α may be involved in human cervical carcinoma dedifferentiation/progression, supporting an association of proline‐directed protein kinase with neoplastic transformation and tumorigenesis. Since protein kinase FA/GSK‐3α may function as a possible regulator of transcription factors/proto‐oncogenes, the results further suggest that kinase FA/GSK‐3α may play a potential role in human cervical carcinogenesis, especially in its dedifferentiation and progression. © 1995 Wiley‐Liss, Inc.
Preferential suppression of insulin‐stimulated proliferation of cultured hepatocytes by somatostatin: Evidence for receptor‐mediated growth regulation
Authors: Kothary Piyush C. , Kokudo Norihiro , Eckhauser F. E. , Delvalle John , Raper Steven E.
Published: May 2005

The role of somatostatin (SS‐14) in the regulation of rat liver regeneration was examined by using thymidine incorporation into hepatocyte DNA labeled with tritiated thymidine, a nuclear‐labeling index, and the binding of 125I‐tyr11‐SS‐14 to hepatocytes isolated at various times after partial hepatectomy. The data demonstrated no suppressive effect of SS‐14 on insulin and glucagon‐stimulated thymidine incorporation into hepatocyte DNA as early as 2 h after partial hepatectomy. These data were substantiated by a nuclear labeling index studies. At 2 h, 125I‐tyr11‐SS‐14 binding to its specific sites on isolated hepatocytes was undetectable. There was a time‐dependent increase in binding of 125I‐tyr11‐SS‐14 to hepatocytes obtained at various times after partial hepatectomy. There was a significant decrease in the number of binding sites after partial hepatectomy as determined by Scatchard analysis. The data were supported by autoradiography analysis of affinity labeled 125I‐tyr11‐SS‐14‐binding protein complex followed by SDS‐PAGE. SS‐14 also inhibited intracellular cAMP in hepatocytes obtained at 18 h after hepatectomy. The data are consistent with the hypothesis that SS‐14 participates via its own receptor in the regulation of the liver regeneration. © 1995 Wiley‐Liss, Inc.
Cytokine induction of proliferation and expression of CDC2 and cyclin a in FDC‐P1 myeloid hematopoietic progenitor cells: Regulation of ubiquitous and cell cycle‐dependent histone gene transcription factors
Authors: Shakoori A. R. , van Wijnen A. J. , Cooper C. , Aziz F. , Birnbaum M. , Reddy G. P. V. , Grana X. , De Luca A. , Giordano A. , Lian J. B. , Stein J. L. , Quesenberry P. , Stein G. S.
Published: May 2005

To evaluate transcriptional mechanisms during cytokine induction of myeloid progenitor cell proliferation, we examined the expression and activity of transcription factors that control cell cycle‐dependent histone genes in interleukin‐3 (IL‐3)‐dependent FDC‐P1 cells. Histone genes are transcriptionally upregulated in response to a series of cellular regulatory signals that mediate competency for cell cycle progression at the G1/S‐phase transition. We therefore focused on factors that are functionally related to activity of the principal cell cycle progression at the G1/S‐phase transition. We therefore focused on factors that are functionally related to activity of the principal cell cycle regulatory element of the histone H4 promoter:CDC2, cyclin A, as well as RB‐and IRF‐related proteins. Comparisons were made with activities of ubiquitous transcription factors that influence a broad spectrum of promoters independent of proliferation or expression of tissue‐specific phenotypic properties. Northern blot analysis indicates that cellular levels of cyclin A and CDC2 mRNAs increase when DNA synthesis and H4 gene expression are initiated, supporting invoulvement in cell cycle progression. Using gel‐shift assays, incorporating factor‐specific antibody and oligonucleotide competition controls, we define three sequential periods following cytokine stimulation of FDC‐P1 cells when selective upregulation of a subset of transcription factors is observed. In the initial period, the levels of SP1 and HiNF‐P are moderately elevated; ATF, AP‐1, and HiNF‐M/IRF‐2 are maximal during the second period; while E2F and HiNF‐D, which contain cyclin A as a component, predominate during the third period, coinciding with maximal H4 gene expression and DNA synthesis. Differential regulation of H4 gene transcription factors following growth stimulation is consistent with a principal role of histone gene promoter elements in integrating cues from multiple signaling pathways that control cell cycle induction and progression. Regulation of transcription factors controlling histone gene promoter activity within the context of a staged cascade of responsiveness to cyclins and other physiological mediators of proliferation in FDC‐P1 cells provides a paradigm for experimentally addressing interdependent cell cycle and cell growth parameters that are operative in hematopoietic stem cells. © 1995 Wiley‐Liss, Inc.
The Rb2/p130 gene product is a nuclear protein whose phosphorylation is cycle regulated
Authors: Baldi Alfonso , De Luca Antonio , Claudio Pier Paolo , Baldi Feliciano , Giordano Giovan Giacomo , Tommasino Massimo , Paggi Marco Giorgio , Giordano Antonio
Published: May 2005

The Rb2/p130 protein has been shown to have a high sequence homology with the retinoblastoma gene product (pRb), one of the most well‐characterized tumor suppressor genes, and with pRb‐related p107, especially in their conserved pocket domains, which display a primary role in the function of these proteins. In this study, we report on the biochemical and immunocytochemical characterization of the Rb2/p130 protein, using a polyclonal antibody developed against its “spacer” region included in the pocket domain of the whole protein. We show that pRb/p130 is a phosphoprotein located at the nuclear level and that its phosphorylation pathway can be dramatically reduced by phosphatase treatment. Moreover pRb/p130, with p107, with p107, is one of the major targets of the E1A viral oncoprotein‐associated kinase activity, showing a phosphorylation pattern which is modulated during the cell cycle, reaching a peak of activation at the onset of S‐phase. © 1995 Wiley‐Liss, Inc.