HORMONAL REGULATION OF MEMBRANE PHENOTYPE IN HEPATOMA CELLS *

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/75361/1/j.1749-6632.1980.tb29527.x.pd

Isolation of rat hepatoma cell variants selectively resistant to dexamethasone inhibition of plasminogen activator

Glucocorticoids induce several phenotypic changes in rat hepatoma cells in tissue culture, including the inhibition of plasminogen activator activity. Variant cell lines resistant to dexamethasone inhibtion of plasminogen activator activity have been isolated using an agar-fibrin overlay technique to identify colonies with fibrinolytic (plasminogen activator) activity. The variants are resistant to concentrations of dexamethasone 1,000 times that necessary to completely inhibit plasminogen activator activity in wild-type cells. The variant phenotype has been inherited in a stable manner for more than 300 generations in continuous culture in the absence of dexamethasone. These variants are unique in that the resistance is not secondary to defective or absent glucocorticoid receptors but is due to a lesion specific for regulation of plasminogen activator. Fluctuation analyses support the hypothesis that resistance to dexamethasone arises randomly and is not induced by dexamethasone. Because HTC cells are heteroploid and karyotypically highly variable, variants are thought to arise primarily by chromosomal segregation events. These variants provide a valuable tool for studying the mechanism of hormonal regulation of plasminogen activator as well as the role of proteases in hormonal regulation of membrane functions.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/49873/1/1040990308_ftp.pd

Cloning and sequencing of cDNA for the rat plasminogen activator inhibitor-1

A cDNA encoding rat plasminogen activator-inhibitor (PAI-1) has been isolated from an HTC rat hepatoma cell cDNA library constructed in phage [lambda]gt10. The cDNA contains 118 bp of 5'-untranslated sequence, 1206 bp encoding a 402-amino acid (aa) protein and 1747 bp of 3'-untranslated sequence. The protein-coding sequence and the derived amino acid sequence share 82% and 81% identity, respectively, with human PAI-1 cDNA and protein. The rat cDNA encodes a preprotein with a 23-aa leader peptide and a predicted N-terminal serine for the mature protein. Three of four potential N-glycosylation acceptor sites as well as the active site of rat PAI-1 are identical to the human protein. The 3'-untranslated region contains a number of unusual regions, including 80 bp of tandemly repeated GpA dinucleotides, a 115-bp stretch which shares greater than 90% sequence identity with a region within the 3'-untranslated cDNA of human PAI-1, and two 70-bp stretches of highly T-rich sequence located close to the 3'-terminus of the cDNA.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/27017/1/0000003.pd

Clinical Use of the Surgeon General’s “My Family Health Portrait” (MFHP) Tool: Opinions of Future Health Care Providers

This study examined medical students’ and house officers’ opinions about the Surgeon General’s “My Family Health Portrait” (MFHP) tool. Participants used the tool and were surveyed about tool mechanics, potential clinical uses, and barriers. None of the 97 participants had previously used this tool. The average time to enter a family history was 15 min (range 3 to 45 min). Participants agreed or strongly agreed that the MFHP tool is understandable (98%), easy to use (93%), and suitable for general public use (84%). Sixty‐seven percent would encourage their patients to use the tool; 39% would ensure staff assistance. Participants would use the tool to identify patients at increased risk for disease (86%), record family history in the medical chart (84%), recommend preventive health behaviors (80%), and refer to genetics services (72%). Concerns about use of the tool included patient access, information accuracy, technical challenges, and the need for physician education on interpreting family history information.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/147053/1/jgc40510.pd

Group 13 HOX proteins interact with the MH2 domain of R-Smads and modulate Smad transcriptional activation functions independent of HOX DNA-binding capability

Interactions with co-factors provide a means by which HOX proteins exert specificity. To identify candidate protein interactors of HOXA13, we created and screened an E11.5–E12.5, distal limb bud yeast two-hybrid prey library. Among the interactors, we isolated the BMP-signaling effector Smad5, which interacted with the paralogous HOXD13 but not with HOXA11 or HOXA9, revealing unique interaction capabilities of the AbdB-like HOX proteins. Using deletion mutants, we determined that the MH2 domain of Smad5 is necessary for HOXA13 interaction. This is the first report demonstrating an interaction between HOX proteins and the MH2 domain of Smad proteins. HOXA13 and HOXD13 also bind to other BMP and TGF-β/Activin-regulated Smad proteins including Smad1 and Smad2, but not Smad4. Furthermore, HOXD13 could be co-immunoprecipitated with Smad1 from cells. Expression of HOXA13, HOXD13 or a HOXD13 homeodomain mutant (HOXD13(IQN>AAA)) antagonized TGF-β-stimulated transcriptional activation of the pAdtrack-3TP-Lux reporter vector in Mv1Lu cells as well as the Smad3/Smad4-activated pTRS(6)-E1b promoter in Hep3B cells. Finally, using mammalian one-hybrid assay, we show that transcriptional activation by a GAL4/Smad3-C-terminus fusion protein is specifically inhibited by HOXA13. Our results identify a new co-factor for HOX group 13 proteins and suggest that HOX proteins may modulate Smad-mediated transcriptional activity through protein–protein interactions without the requirement for HOX monomeric DNA-binding capability

Strategies to Prevent Anthracycline-Related Congestive Heart Failure in Survivors of Childhood Cancer

Cardiovascular complications are a leading cause of therapy-related morbidity and mortality in long-term survivors of childhood malignancy. In fact, childhood cancer survivors are at a 15-fold risk of developing CHF compared to age-matched controls. There is a strong dose-dependent association between anthracycline exposure and risk of CHF, and the incidence increases with longer followup. Outcome following diagnosis of CHF is generally poor, with overall survival less than 50% at 5 years. The growing number of childhood cancer survivors makes it imperative that strategies be developed to prevent symptomatic heart disease in this vulnerable population. We present here an overview of the current state of knowledge regarding primary, secondary, and tertiary prevention strategies for childhood cancer survivors at high risk for CHF, drawing on lessons learned from prevention studies in nononcology populations as well as from the more limited experience in cancer survivors

Paradoxical effects of glucocorticoids on regulation of plasminogen activator activity : Mediation by glucocorticoid receptors

Dexamethasone, a synthetic glucocorticoid, decreases the plasminogen activator (PA) activity of HTC rat hepatoma cells in tissue culture. Paradoxically, dexamethasone enhances the cyclic nucleotide stimulation of PA activity in these cells 2-4-fold. In this report, we investigated whether this paradoxical glucocorticoid effect is mediated by the same proximal events which mediate such direct regulatory actions of glucocorticoids as the induction of tyrosine aminotransferase activity. We compared the concentration-dependences for several classes of steroids, previously classified as full agonists, partial agonists, antagonists or inactive steroids with respect to induction of the transaminase, for both enhancement of cyclic nucleotide stimulation of PA activity and induction of tyrosine aminotransferase activity in parallel cultures. The full agonists dexamethasone and cortisol, the partial agonists deoxycorticosterone and 11[beta]-hydroxyprogesterone, the inactive steroid tetrahydrocortisol, and the antagonist 17[alpha]-methyltestosterone exhibited similar potencies with respect to both phenomena. Furthermore, when cells were incubated with both dexamethasone and 17[alpha]-methyltestosterone, the latter blocked enhancement by dexamethasone in a concentration-dependent fashion. We conclude that glucocorticoid enhancement of cyclic nucleotide stimulation of PA activity is mediated by the same glucocorticoid receptors which mediate direct regulatory effects.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/24927/1/0000354.pd

Strategies to prevent anthracycline-related congestive heart failure in survivors of childhood

Cardiovascular complications are a leading cause of therapy-related morbidity and mortality in long-term survivors of childhood malignancy. In fact, childhood cancer survivors are at a 15-fold risk of developing CHF compared to age-matched controls. There is a strong dose-dependent association between anthracycline exposure and risk of CHF, and the incidence increases with longer followup. Outcome following diagnosis of CHF is generally poor, with overall survival less than 50% at 5 years. The growing number of childhood cancer survivors makes it imperative that strategies be developed to prevent symptomatic heart disease in this vulnerable population. We present here an overview of the current state of knowledge regarding primary, secondary, and tertiary prevention strategies for childhood cancer survivors at high risk for CHF, drawing on lessons learned from prevention studies in nononcology populations as well as from the more limited experience in cancer survivors

Hormonal regulation of plasminogen activator in rat hepatoma cells

Plasminogen activators are membrane-associated, arginine-specific serine proteases which convert the inactive plasma zymogen plasminogen to plasmin, an active, broad-spectrum serine protease. Plasmin, the major fibrinolytic enzyme in blood, also participates in a number of physiologic functions involving protein processing and tissue remodelling, and may play an important role in tumor invasion and metastasis. In HTC rat hepatoma cells in. tissue culture, glucocorticoids rapidly decrease plasminogen activator (PA) activity. We have shown that this decrease is mediated by induction of a soluble inhibitor of PA activity rather than modulation of the amount of PA. The hormonally-induced inhibitor is a cellular product which specifically inhibits PA but not plasmin. We have isolated variant lines of HTC cells which are selectively resistant to the glucocorticoid inhibition of PA but retain other glucocorticoid responses. These variants lack the hormonally-induced inhibitor; PA from these variants is fully sensitive to inhibition by inhibitor from steroid-treated wild-type cells. Cyclic nucleotides dramatically stimulate PA activity in HTC cells in a time- and concentration-dependent manner. Paradoxically, glucocorticoids further enhance this stimulation. Thus glucocorticoids exert two separate and opposite effects on PA activity. The availability of glucocorticoid-resistant variant cell lines, together with the unique regulatory interactions of steroids and cyclic nucleotides, make HTC cells a useful experimental system in which to study the multihormonal regulation of plasminogen activator.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/45346/1/11010_2004_Article_BF00225243.pd