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Biomedical subjects

M D Johnson

Publications and source records attributed to M D Johnson.

At least 127 records · Page 7Linked to original sources
PubMed1995

The young female athlete.

It is important that girls and young women participate in sports and develop skills that promote lifelong athletic participation, because of the psychological, sociologic, and physiologic benefits associated with exercise. When an athlete begins intensive, competitive exercise training at a young age, or when the preoccupation with thinness supersedes a desire to be healthy, potential morbidity results. Lack of information and the strong desire to win contribute to this problem. There is relatively little known about the long-term physical and psychological effects of early intensive athletic training and the female athlete triad on the young female athlete. In addition to the need for further research in these areas, there is a need for education of physicians, coaches, trainers, athletes, and parents. The preparticipation physical examination is an excellent opportunity for the physician to screen for the triad disorders and educate athletes and parents on healthy nutrition, normal menstrual function, and the benefits of exercise.

Adolescent↗
PubMed1995

Synergistic interaction of transforming growth factor alpha and c-myc in mouse mammary and salivary gland tumorigenesis.

The c-myc oncogene is commonly amplified in breast cancer and is known to interact synergistically with transforming growth factor alpha (TGF alpha) in vitro to promote phenotypic transformation of mammary epithelial cells. In addition, both genes are under sex steroid hormone regulation in breast cancer. We have used a bitransgenic mouse approach to test the relevance of Myc-TGF alpha interaction in mammary gland tumorigenesis of virgin animals in vivo. We mated single transgenic TGF alpha and c-myc mouse strains to yield double transgenic offspring for TGF alpha and c-myc. All (20 of 20) double transgenic TGF alpha/c-myc animals developed synchronous mammary tumors at a mean age of 66 days. An unexpected finding was that tumor latency and frequency in males and virgin females were identical. Thus, two gene products that are known to be coinduced in breast cancer by the sex hormones estrogen and progesterone strongly synergize to induce synchronous mammary tumors, independent of sex. The tumors, despite being estrogen receptor positive, were readily transplanted as highly malignant s.c. cancers in ovariectomized nude mice. Although approximately one-half of single transgenic c-myc virgin females also eventually developed mammary gland tumors, these were stochastic and arose after a long latency period of 9-12 months. Single transgenic virgin TGF alpha females and males, c-myc males, and transgene-negative littermates did not develop tumors (ages up to 15 months). The salivary glands of double transgenic animals also coexpress the two transgenes and show pathological abnormalities ranging from hyperplasias to frank adenocarcinomas. In contrast, the salivary glands of single transgenic and wild-type animals showed only mild hyperplasias or metaplasias, but tumors were not observed. In situ hybridization analysis of mammary and salivary glands revealed that hyperplastic and tumorous areas colocalize with regions that overexpress both the TGF alpha and c-myc transgenes. This indicates that there is a requirement for the presence of both proteins for transformation of these glands. In summary, TGF alpha and c-Myc synergize in an extremely powerful way to cause breast and salivary gland tumorigenesis in males and virgin females without a requirement for pregnancies.

Adenocarcinoma↗
PubMed1995

Beta-adrenoceptor-G alpha S coupling decreases with age in rat aorta.

beta-Adrenoceptor (beta AR) responsiveness, receptor density, receptor-G protein coupling, and the possible role of membrane fluidity in receptor-G protein coupling were investigated in the rat aorta with age. The beta AR agonist isoproterenol (ISO) produced relaxation of KCl-induced aortic contractions by 97%, 21%, and 0% in aortae from 1- 6-, and 24-month-old Fischer 344 rats, respectively. Forskolin completely relaxed the contractions at all ages. beta AR density was determined in aortic membranes by saturation binding of 125I-cyanopindolol (125I-CYP). beta AR density was 76, 52, and 47 fmol/mg of protein in 1-, 6-, and 24-month-old rats, respectively. To investigate beta AR coupling to G proteins, displacement by ISO of 125I-CYP binding was determined in aortic membranes in the presence and absence of the GTP analog guanosine-5'-(beta gamma-imido)triphosphate [Gpp(NH)p] (0.1 mM). The effect of Gpp(NH)p on the ISO displacement curve for 125I-CYP binding was greatest in 1-month-old rats and decreased markedly with age. In 1-month-old aorta, in the absence of Gpp(NH)p the ISO displacement curve was biphasic and two affinity constants were determined (KH - 0.061 microM and KL = 2.4 microM). In the presence of Gpp(NH)p the ISO displacement curve was monophasic (Kd - 0.72 microM). In 6-month-old aorta, whereas an effect of Gpp(NH)p on the ISO displacement curve could still be observed [in the absence of Gpp(NH)p, KH = 0.2 microM and KL = 3.5 microM; in the presence of Gpp(NH)p, Kd - 0.83 microM], the affinity constant for high affinity agonist binding and the percentage of receptors with high affinity for agonist were decreased significantly. In 24-month-old aorta there was no effect of Gpp(NH)p on the ISO displacement curve and a single affinity constant was detected [0.7 microM and 0.8 microM in the presence and absence of Gpp(NH)p, respectively]. The presence of two affinity constants for ISO in 1- and 6-month-old aorta in the absence of Gpp(NH)p and single affinity constants in the presence of Gpp(NH)p presumably represent the G protein-coupled and uncoupled states of the beta ARs, which are not observed in 24-month-old aorta. The ability of the beta AR to form the high affinity nucleotide-sensitive complex with the agonist was restored by treatment of the membranes with cis-vaccenic acid, which increases the fluidity of the membrane.(ABSTRACT TRUNCATED AT 400 WORDS)

Aging↗
PubMed1995

The expression of alpha 1 adrenoceptor subtypes changes with age in the rat aorta.

Previous studies showed that alpha 1 adrenoceptor-mediated contractile responses change with age in the rat aorta, becoming more sensitive to Ca++ channel blockers and less sensitive to chlorethylclonidine (CEC), suggesting a change in the alpha 1 adrenoceptor subtypes that are present. In this study, alpha 1 adrenoceptor density and alpha 1 adrenoceptor subtypes were measured in the Fischer 344 rat aorta during aging. Aortic alpha 1 adrenoceptor densities, determined by saturation binding of 2-[beta-(4-hydroxy-3-[125I]iodophenyl)ethylaminomethyl] tetralone ([125I]-HEAT), were 47, 41 and 45 fmol/mg protein in 1-,6- and 24-month-old rats, respectively. The noncompetitive antagonist CEC completely blocked [125I]-HEAT binding in aortas from 1-month-old rats but inhibited binding only partially in aortas from older rats. Two binding sites were detected for norepinephrine and for WB4101 in all ages. The low-affinity constants for WB4101 (31-51 nM) were consistent with those for the alpha 1b adrenoceptor subtype, and this binding site decreased with age. The high-affinity constant for WB4101 (1.4 nM) in 1-month-old aorta was consistent with that for alpha 1d adrenoceptor subtype, whereas the high-affinity constants (0.03 nM) in 6- and 24-month-old aortas were consistent with those for the alpha 1a adrenoceptor subtype. At least three alpha 1 adrenoceptor subtypes appear to be colocalized in the rat aorta, so the binding affinities may reflect binding to more than one subtype. This makes it difficult to identify denfinitively the subtypes based on their radioligand binding characteristics.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗
PubMed1994

Age-related changes in angiotensin II-stimulated vascular contraction and inositol phosphate accumulation in Fischer 344 rats.

This study examined the influence of age on angiotensin II (AII)-stimulated vascular contractile responses and inositol phosphate (IP) accumulation in Fischer 344 rats. In the aorta, AII-stimulated contraction and IP accumulation were markedly reduced in 6- and 24-month-old rats compared to 1-month-old rats. There was not a significant difference in the contractile response to AII between 6- and 24-month-old rats, although IP hydrolysis showed a further decrease between 6 and 24 months. In tail artery, there were no differences in contraction and phosphoinositol metabolism in response to AII in the different ages. Losartan blocked AII-stimulated vascular contraction and IP hydrolysis in both aorta and tail artery while PD123319 did not inhibit either response. These data indicate that during maturation, there is a decline in AII-stimulated aortic contraction and IP accumulation in aorta but not in tail artery and these changes are due to altered AT1 receptor function.

Aging↗
PubMed1994

Targeted disruption of the Hexa gene results in mice with biochemical and pathologic features of Tay-Sachs disease.

Tay-Sachs disease, the prototype of the GM2 gangliosidoses, is a catastrophic neurodegenerative disorder of infancy. The disease is caused by mutations in the HEXA gene resulting in an absence of the lysosomal enzyme, beta-hexosaminidase A. As a consequence of the enzyme deficiency, GM2 ganglioside accumulates progressively, beginning early in fetal life, to excessive amounts in the central nervous system. Rapid mental and motor deterioration starting in the first year of life leads to death by 2-4 years of age. Through the targeted disruption of the mouse Hexa gene in embryonic stem cells, we have produced mice with biochemical and neuropathologic features of Tay-Sachs disease. The mutant mice displayed < 1% of normal beta-hexosaminidase A activity and accumulated GM2 ganglioside in their central nervous system in an age-dependent manner. The accumulated ganglioside was stored in neurons as membranous cytoplasmic bodies characteristically found in the neurons of Tay-Sachs disease patients. At 3-5 months of age, the mutant mice showed no apparent defects in motor or memory function. These beta-hexosaminidase A-deficient mice should be useful for devising strategies to introduce functional enzyme and genes into the central nervous system. This model may also be valuable for studying the biochemical and pathologic changes occurring during the course of the disease.

Animals↗
PubMed1994

Isoform specific interactions of troponin I and troponin C determine pH sensitivity of myofibrillar Ca2+ activation.

We investigated whether differences in isoforms of troponin I (TnI) and troponin C (TnC) can account for the greater inhibition of Ca(2+)-dependent MgATPase activity by acidic pH in cardiac (c) than in fast skeletal (fs) myofilaments. We studied fast skeletal myofibrils from which whole Tn was extracted by displacement with excess fsTnT (the tropomyosin binding subunit of Tn) followed by reconstitution with TnC-TnI. Exchange of fsTnI with cTnI did not alter the effect of a drop in pH from 7.0 to 6.5 on the relation between pCa (-log[Ca2+]) and MgATPase activity of fast skeletal myofibrils. Exchange of fsTnC with cTnC did, however, induce an increase in the effect of this same pH change on Ca2+ activation. Yet, the pH sensitivity of Ca2+ activation of fast skeletal myofibrils containing cTnC was not as great as that of native cardiac myofibrils. However, when both fsTnC and fsTnI of fast skeletal myofibrils were replaced by cTnC-cTnI, there was a pH-induced shift in Ca2+ sensitivity similar to that of cardiac myofibrils. In studies using fluorescent probes, both pure fsTnC and pure cTnC showed decreased Ca2+ binding as pH was lowered. This decrease was potentiated in the fsTnC-fsTnI and cTnC-cTnI complexes. However, the effect of acidic pH was the same in fsTnC and the hybrid complex, fsTnC-cTnI, and in cTnC and the hybrid complex, cTnC-fsTnI. Thus, isoform specific interactions between TnI and TnC appear important in the differential response of skeletal and cardiac myofilaments to acidosis.

Animals↗
PubMed1994

Inhibition of angiogenesis by tissue inhibitor of metalloproteinase.

Matrix proteases play a critical role in cell invasion and migration, including the process of angiogenesis. The ability of specific factors to induce angiogenic responses correlates with their stimulation of matrix protease synthesis and release. Using an in vivo angiogenesis assay, the endothelial cell response to known angiogenic factors, basic fibroblast growth factor (bFGF) and adipocyte conditioned medium, was blocked by an inhibitor of matrix metalloproteinase activity, TIMP-1. The TIMP effect was mediated, at least in part, through the inhibition of endothelial cell migration, as determined by the ability of TIMP to block chemotaxis in a Boyden chamber assay. These results indicate that the inhibition of migration is a direct effect on the endothelial cells and does not require accessory cells. An additional observation was that the RNA levels for TIMP were significantly reduced in differentiated adipocytes, compared to undifferentiated F442A controls. Therefore, the acquisition of an angiogenic phenotype may involve not only the induction of positive factors, but also the suppression of angiogenesis inhibitors.

Adipocytes↗
PubMed1994

Solubilization of a tripeptide HIV protease inhibitor using a combination of ionization and complexation with chemically modified cyclodextrins.

Kynostatin (KNI-272), an experimental HIV protease inhibitor, is currently undergoing preclinical testing for the treatment of AIDS. This transition state mimetic tripeptide exhibits extremely low aqueous solubility (4 micrograms/mL) making target concentrations (5-50 mg/mL) for parenteral solution formulations difficult to achieve. The presence of an ionizable (5-isoquinolinyloxy)acetyl moiety makes solubilization via pH adjustment possible, but a solubility > 5 mg/mL requires an adjustment in pH below 2.0, which would be physiologically unacceptable. This study examines and compares two approaches for solubilizing kynostatin: (1) inclusion complex formation at chemically distinct hydrophobic binding sites using (2-hydroxypropyl)-beta-cyclodextrin (HPCD) and a sulfobutyl ether derivative of beta-cyclodextrin (beta-CD-SBE) and (2) a combined strategy utilizing ionization of the isoquinoline moiety coupled with inclusion complex formation at the remaining binding site(s). Macroscopic binding constants determined from solubility profiles as a function of pH and HPCD concentration have been compared with the microscopic binding constant for formation of the isoquinoline-HPCD inclusion complex determined by UV difference spectroscopy to examine the independence of binding domains within KNI-272. As demonstrated in this report, combination strategies tailored to the properties of different domains within the molecule may be highly effective in solubilizing compounds such as poorly soluble peptides.

Chromatography, High Pressure Liquid↗
PubMed1994

Molecular cloning of TPAR1, a gene whose expression is repressed by the tumor promoter 12-O-tetradecanoylphorbol 13-acetate (TPA).

We previously isolated a partial cDNA sequence, termed TPAR1 (TPA repressed gene 1), from a cDNA library constructed from C3H10T1/2 mouse embryo fibroblasts treated with TPA, using a differential screening procedure. (M.D. Johnson et al. Mol. Cell. Biol. 7, 2821-2829, 1987). In the present study, we have cloned two corresponding full-length 1.9- and 3.4-kb cDNAs of TPAR1 from murine cDNA libraries. Sequence analysis of these TPAR1 cDNAs revealed that they encode 89 and 93 amino acid polypeptides, respectively, with a putative leader sequence and show significant homology with the human cytokine interleukin-8 (IL-8) and its superfamily. Genomic DNA isolation and structural characterization provide evidence that the TPAR1 mRNAs are transcribed from a single gene with alternative splicing. TPAR1 mRNAs are expressed ubiquitously among adult mouse tissues as three major transcripts, 1.9, 3.4, and 6.5 kb, whose expression depends on the tissue type. The levels of TPAR1 mRNAs were markedly decreased in fibroblasts following TPA treatment and also in serum-deprived quiescent fibroblasts stimulated by serum. The levels of TPAR1 mRNAs were dramatically down-regulated in regenerating rat liver when compared to normal adult liver. In addition, there was no detectable expression of TPAR1 in three rat hepatoma cell lines and several transformed fibroblast cell lines. Thus, the TPAR1 gene is a new member of the cytokine IL-8 superfamily, whose expression is down-regulated in rapidly dividing cells. Further studies are required to determine whether it plays a negative role in controlling cell proliferation and tumorigenesis.

Amino Acid Sequence↗
PubMed1994

Matrix-degrading proteases in hormone-dependent breast cancer.

Proteases have emerged as important modulators of the metastatic capacity of cancer. However, metastasis is regulated by multiple other characteristics of the tumor cell and evidence suggests the participation of multiple classes of proteases. In the present article we review the literature concerning the potential biological roles of multiple proteases in breast cancer. In particular, we focus on the gelatin-degrading metallo proteinase and on a novel 80 KDa matrix-degrading protease that appears to be commonly expressed in hormone dependent breast cancer cell lines.

Adenocarcinoma↗
PubMed1994

Prenatal cocaine exposure affects the development of aortic adrenergic innervation and contractile responses.

This study examines the effects of prenatal cocaine administration on the development of vascular sympathetic innervation and contractile responsiveness. Rabbits received cocaine (4 mg/kg, iv, bid) or saline during gestational days 8 to 29. Aortas were obtained on postnatal days 10, 20, 30 and 50. Vascular smooth muscle responsiveness was assessed by measuring aortic contractile responses to norepinephrine (NE) and to other vasoconstrictors. Vascular adrenergic innervation was evaluated by measuring desipramine sensitive [3H]-NE uptake into aortic ring segments and aortic NE content. [3H]-NE uptake and NE content were reduced at postnatal days 10 and 20 in the rabbits exposed prenatally to cocaine. Differences were not observed at postnatal days 30 or 50. The contractile response to NE was reduced in rabbits exposed to cocaine prenatally. Maximal response and potency were decreased at postnatal day 10 and potency was still decreased at day 20, but not at the older ages. Contractile responses to serotonin (5-HT) and angiotensin II (AII) were not affected by prenatal cocaine exposure. These results suggest that prenatal cocaine exposure delays the development of aortic adrenergic innervation and alpha adrenoceptor responsiveness.

Angiotensin II↗
PubMed1994

The epidermal growth factor receptor is associated with phospholipase C-gamma 1 in meningiomas.

In this study we evaluated phospholipase C-gamma 1 (PLC-gamma 1) expression, activity, and association with the epidermal growth factor (EGF) receptor in a series of human meningiomas as well as cultured meningioma cells. Phospholipase C-gamma 1 was detectable by immunoblot and immunohistochemistry in 13 of 13 meningioma specimens. Epidermal growth factor receptors were detected by immunoblot in six of nine meningiomas (67%) and by immunohistochemistry in 13 of 19 meningiomas (68%) but not in normal leptomeningeal cells. In two of three meningiomas EGF receptors and/or a 170-kd phosphotyrosine band precipitated with a PLC-gamma 1 antiserum. Both PLC-gamma 1 and EGF receptors also exhibited the same pattern of immunostaining on meningioma tissue sections. Phospholipase C-gamma 1 catalytic activity, measured in a PIP2 hydrolysis assay, was higher in nine EGF receptor-positive meningiomas than in six EGF receptor-negative meningiomas (P = .05; t test). Finally, treatment of cultured meningioma cells with transforming growth factor-alpha induced a 78% increase in PLC-gamma 1 catalytic activity. Thus, these data are consistent with the possibility that the EGF receptor tyrosine kinase regulates PLC-gamma 1 activity in native meningioma tissue.

Adult↗
PubMed1994

Electrophysiological and histochemical properties of postnatal rat serotonergic neurons in dissociated cell culture.

Serotonin modulates a variety of neural processes, and is present in a subpopulation of neurons in the raphe nuclei. To study their electrophysiological properties, cells from the mesopontine raphe nuclei of the neonatal rat were dissociated and grown for up to 10 weeks in microcultures. Approximately one third of the neurons were identified as serotonergic based on the presence of serotonin immunoreactivity, tryptophan hydroxylase immunoreactivity, or a high affinity monoamine transporter. About 5% of cultured raphe neurons contained tyrosine hydroxylase immunoreactivity, while 25% contained GABA immunoreactivity. However, no neurons contained both serotonin and tyrosine hydroxylase staining, and less than 1% displayed both serotonin and GABA immunoreactivities. Cultured serotonergic neurons did not exhibit pacemaker firing in the presence of alpha 1 adrenergic receptor agonists such as phenylephrine or norepinephrine. Approximately one third were hyperpolarized by serotonin or the selective serotonin1A receptor agonist, (+/-)-8-hydroxy-2-(di-N-propylamino)tetralin. Virtually all serotonergic neurons responded to application of glutamate, kainate, N-methyl-D-aspartate, GABA, and glycine. Depolarizing and hyperpolarizing synaptic potentials blocked by glutamate or GABAA receptor antagonists were frequently observed in both serotonergic and non-serotonergic raphe neurons. Slow inhibitory postsynaptic potentials were evoked by activating single presynaptic serotonergic neurons with a brief intracellular current pulse. The slow inhibitory synaptic potential had a mean latency to onset of 35 +/- 5 ms, a duration of 0.8-2.6 s, and was inhibited by the serotonin1A autoreceptor antagonists, (-)propranolol and spiperone. The rising and falling phases of the inhibitory potential could be fit by single exponential functions with mean time constants of 53 +/- 8 ms and 504 +/- 78 ms, respectively. Serotonin1A receptor-mediated autoinhibition was observed in microcultures containing a solitary serotonergic neuron, and thus constituted synaptic serotonin release, responsiveness, and re-uptake by a single vertebrate neuron. In summary, histochemical and electrophysiological evidence was obtained for catecholaminergic, GABAergic, and glutamatergic non-serotonergic raphe neurons in culture, many of which formed functional synaptic connections with neighboring cells. Additionally, cultured mesopontine serotonergic neurons expressed many of the cytochemical markers, neurotransmitter receptors, and synaptic functions observed in such cells in vivo, but the proportion of neurons sensitive to serotonergic and adrenergic agonists was significantly less than that reported in vivo. For the first time, the kinetics and pharmacology of serotonergic synaptic transmission by a single vertebrate serotonergic raphe neuron were determined, and found to resemble those observed after extracellular stimulation of populations of raphe neurons in slices and in vivo.

5,7-Dihydroxytryptamine↗
PubMed1994

Synaptic glutamate release by postnatal rat serotonergic neurons in microculture.

Serotonergic neurons are thought to play a role in depression and obsessive compulsive disorder. However, their functional transmitter repertoire is incompletely known. To investigate this repertoire, intracellular recordings were obtained from 132 cytochemically identified rat mesopontine serotonergic neurons that had re-established synapses in microcultures. Approximately 60% of the neurons evoked excitatory glutamatergic potentials in themselves or in target neurons. Glutamatergic transmission was frequently observed in microcultures containing a solitary serotonergic neuron. Evidence for co-release of serotonin and glutamate from single raphe neurons was also obtained. However, evidence for gamma-aminobutyric acid release by serotonergic neurons was observed in only two cases. These findings indicate that many cultured serotonergic neurons form glutamatergic synapses and may explain several observations in slices and in vivo.

Animals↗