Our previous finding that insulin induces apolipoprotein AI (apoAI) transcription points to the participation of intracellular signaling. This finding prompted us to ask whether two classical G-protein-coupled signaling pathways requiring activated protein kinase A (PKA) or kinase C (PKC) may also regulate apoAI. Therefore, human hepatoma, Hep G2 cells stably transfected with pAI.474-CAT, a reporter construct spanning -474 to -7 of apoAI DNA fused to chloramphenicol acetyltransferase (CAT) were treated with 10 microm forskolin (FSK) or 50 nm phorbol dibutyrate (PDBu) to activate PKA and PKC, respectively. Results showed that the apoAI promoter activity increased 4-5-fold following 24 h of treatment with either FSK or PDBu. Induction by either agent was blocked with actinomycin D but not the protein synthesis inhibitor, cycloheximide. The PKA inhibitor, PKI 14-22 amide, abrogated induction by FSK, 100 microm 8-bromo-cAMP, or 100 ng/ml cholera toxin, but it had no effect on activation via PKC. Similarly, PDBu induction was attenuated by 2 microm of the PKC inhibitor, GF109203X, but it did not affect FSK activity. Next we used deletional constructs to show that the actions of FSK and PDBu required the insulin-responsive core element (IRCE). This motif matched the consensus binding site for the transcription factor, Sp1. The binding of Sp1 to the IRCE was confirmed by gel-retardation and supershift analysis. Site-directed mutagenesis of the IRCE eliminated Sp1 action and induction by FSK or PDBu. Whereas overexpression of Sp1 enhanced basal and FSK or PDBu induced promoter activity, transfection of an antisense oligomer against Sp1 mRNA attenuated both parameters. In summary, activation of PKA or PKC increases apoAI promoter activity. The activity of both signaling pathways is mediated by the IRCE, a motif that binds the transcription factor, Sp1.

Apolipoprotein A-I↗

PubMed2000

Midkine promoter-based adenoviral vector gene delivery for pediatric solid tumors.

It is important to develop a system to express therapeutic genes in tumor cells with sufficient selectivity for cancer gene therapy. Midkine (MK) is a newly identified heparin-binding growth factor that is transiently expressed in the early stages of retinoic acid-induced differentiation of embryonal carcinoma cells. It has been reported that many human malignant tumors express high levels of MK mRNA or protein. However, no MK expression is detected in human or mouse liver. These interesting features of MK led us to examine the MK promoter as a candidate for tumor-specific gene expression. We thus developed new recombinant adenoviral (Ad) vectors containing either luciferase reporter gene (AdMKLuc) or herpes simplex thymidine kinase gene (AdMKTK) under the control of the human MK promoter. AdMKLuc achieved relatively high activity in Wilms' tumor (G-401) and neuroblastoma (SK-N-SH) cell lines. In addition, AdMKTK induced marked cell death in response to ganciclovir (GCV) in these same lines. Conversely, very low activity of the MK promoter was observed in mouse liver in vivo compared with the cytomegalovirus promoter. Importantly, AdMKTK + GCV did not induce liver toxicity, whereas substantial toxicity was seen with AdCMVTK + GCV treatment. On the basis of these findings, we conclude that the MK promoter is a candidate tumor-specific promoter for Wilms' tumor or neuroblastoma.

Adenoviridae↗

PubMed2000

Effects of irradiation on the sensitivity of melatonin release to norepinephrine in cultured pineal bodies of rats.

We measured levels of basal and norepinephrine (NE) -induced melatonin release in cultured pineal bodies of rats just after and 2 days after irradiation (20 or 200 Gy) or sham-irradiation. On the day of irradiation, basal and NE-induced melatonin release were not affected by irradiation. Two days after irradiation, basal melatonin levels in both irradiated groups were significantly lower than those in control group, whereas NE-induced melatonin levels in the irradiated groups were higher than those in the control group. These results suggested that irradiation decreases basal melatonin synthesis in pineal glands but that melatonin response to NE is increased after irradiation.

Adrenergic alpha-Agonists↗

PubMed2000

The correlation between P300 alterations and regional cerebral blood flow in non-demented Parkinson's disease.

P300 was evoked by a visual oddball and an S1-S2 task in 22 non-demented Parkinson's disease (PD) patients (13 in the early stage, nine in the late stage) and 18 normal controls. Reaction time was also measured. All patients undertook the (99)Tc-ECD SPECT examination. Quantitative regional cerebral blood flow (rCBF) was obtained by overlying SPECT image on the 3D-magnetic resonance image. In the PD patients in the late stage, P300 latency to S2-same and reaction time were significantly prolonged, while rCBF at bilateral frontal, temporal, and the right parietal lobes was decreased. P300 latency to S2-same was significantly correlated with the rCBF at bilateral temporal lobes. Reaction time was significantly correlated with the rCBF at the right frontal and parietal lobes, as well as the temporal and occipital lobes. The results suggest that P300 changes in non-demented PD in the late stage could be related to the temporal lobe dysfunction.

Aged↗

PubMed2000

Enzyme histochemistry on human placental trophoblasts: the effect of fixation delay on enzyme activity.

We examined the effect of time delay in tissue fixation on enzyme histochemically detectable enzyme activity and its localization in term human placental trophoblasts. Four placental enzymes, alkaline phosphatase, acid phosphatase, glucose-6-phosphatase, and cytochrome c oxidase, were studied. A fixation delay of 15 min did not markedly alter the activity or distribution pattern of the four enzymes, excepted for a slight reduction in cytochrome c oxidase activity and the appearance of dilated endoplasmic reticula positive for glucose-6-phosphatase. A fixation delay of 60 min abolished cytochrome c oxidase activity, but the activities of the other three enzymes remained positive. When the placental tissue was stored at 4 degrees C without cutting for 24 h before fixation, cell degeneration occurred. However, alkaline phosphatase activity was still clearly demonstrable. In enzyme histochemistry, ,,immediate" fixation is superior, but even if this cannot be performed, the placentas, especially when they are from patients with rare disorders, should not be discarded. Observations made here will be useful for clinician's attempting enzyme histochemistry in organs other than the placenta.

Acid Phosphatase↗

PubMed2000

Enzyme-histochemically detectable glucose-6-phosphatase is present in chorion laeve trophoblasts of human fetal membranes.

The purpose of the present study was to demonstrate the presence of glucose-6-phosphatase (G6Pase) in fetal membranes from various gestational ages (20-40 weeks of gestation). Ultrastructural enzyme-histochemical analysis of G6Pase was performed using cerium and lead as capturing agents. Precipitates indicating G6Pase activity were present mainly in the endoplasmic reticulum and partly in the nuclear envelope of chorion laeve trophoblasts, but absent in amniotic epithelial cells. Stringent histochemical control experiments performed ensured specific detection of G6Pase activity. The results indicate that histochemically detectable G6Pase is present in the chorion laeve trophoblasts of human fetal membranes. This enzyme may have some physiological significance in carbohydrate metabolism in human fetal membranes and regulation of amniotic fluid glucose concentration.

Amnion↗

PubMed2000

Meconium-stained amniotic fluid exhibits chemotactic activity for polymorphonuclear leukocytes in vitro.

We have studied whether meconium-stained, turbid amniotic fluid (turbid AF) obtained during term pregnancy possesses chemotactic activity for polymorphonuclear leukocytes (PMNs) in the absence of clinically apparent infection. Eight samples of turbid AF were obtained from eight women who underwent a cesarean section (four emergency and four elective cesarean sections) in the absence of signs of clinical infection or fetal distress. Samples of clear AF obtained from nine women during an elective cesarean section served as a control. We used also a negative control (medium only) and a positive control containing 10 nM N-formyl-methionyl-leucyl-phenylalanine. The control or turbid AF specimen was placed in the lower compartment of a blind well chamber, and the PMN suspension was placed in the upper compartment. Following incubation, the number of PMNs that had migrated and passed through the filter to the AF was counted. The number of control PMNs that migrated to the turbid AF (200+/-59) was comparable to that of the positive (162+/-24) but significantly exceeded that of the clear AF (17+/-11; P < 0.0001) and of the negative control (25+/-9; P < 0.0001). Checkerboard assay indicated that the turbid AF exhibited a dose-dependent chemotactic activity for PMNs. The turbid AF contained higher levels of TNFalpha, IL-1beta and IL-8 than the clear AF. The concentration of IL-8 in the AF was correlated positively with the chemotactic activity of the AF (r = 0.733, P = 0.0005). Anti-human IL-8 antibody added in the turbid AF dose-dependently abolished the chemotactic activity of the turbid AF. It is concluded that meconium-stained AF is a chemoattractant for PMNs and that cytokines such as an IL-8 may be involved in this phenomenon.

Adult↗

PubMed2000

MR in vivo imaging of oxygen suppression effect of soft contact lens on the human cornea.

The O(2) suppression effect of a soft contact lens on the human cornea was measured using dynamic magnetic resonance imaging (MRI) of the anterior chamber transcorneally exposed to O(2). Dynamic T(1)-weighted fast spin echo imaging of anterior chambers (TR = 2 s, TE = 15 ms, 5-mm slice) was performed both before and during oxygen supply to a full goggle placed on the face of volunteers wearing a soft contact lens on one eye and nothing on the other eye as a control. Within 15 min after O(2) administration, significantly lower intensity changes were obtained in the anterior chambers of the eyes with the contact lens than in those of the eyes without one, suggesting that dynamic MRI of the anterior chamber transcorneally exposed to O(2) can be used to evaluate the O(2) suppression effect of a soft contact lens on the cornea.

Anterior Chamber↗

PubMed2000

Release of allergenic proteins from rice grains induced by high hydrostatic pressure.

Protein release from rice grains during high-pressure treatment was investigated. When polished rice grains were immersed in distilled water and pressurized at 100-400 MPa, a considerable amount of proteins (0.2-0.5 mg per gram of grains) was released. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analyses, the major proteins released were identified as 16 kDa albumin, alpha-globulin, and 33 kDa globulin, which were known as major rice allergens. By scanning electron microscopic observation of rice grains pressurized at 300 MPa, partial morphological changes in endosperm cells but no apparent structural changes in protein bodies were detected. The content of these allergenic proteins decreased by pressurization and almost completely disappeared from rice grains by the pressurization in the presence of proteolytic enzyme. These results suggest that partial destruction of endosperm cells caused by pressurization enhances permeability of a surrounding solution into rice grains and that a part of the proteins are solubilized and subsequently released into a surrounding solution.

Allergens↗

PubMed2000

Occult vascular malformations of the spinal cord: report of four cases not detected by angiography.

Angiographically occult vascular malformations (AOVMs) are reported. Although cavernous malformations, the most common type among the spinal intramedullary AOVMs, have been well documented, the natural history and clinical features of non-cavernous type AOVMs of the spinal cord remain obscure. We have evaluated four cases of non-cavernous AOVMs in the past 9 years. Although the clinical features and magnetic-resonance imaging (MRI) of three cases were similar to those previously reported, the other one had unusual features of insidious onset with limb asymmetry. The natural history of intramedullary occult AVMs remains enigmatic. One speculation for the neurological deterioration is that it results from repeated hemorrhages around the vessels or from intralumenal thrombosis.

Adult↗

PubMed2000

Ultracytochemical localization of 5'-nucleotidase activity in human ejaculated spermatozoa.

5'-nucleotidase, an adenosine producing enzyme with a glycosylphosphatidylinositol-anchored structure, was localized in human ejaculated spermatozoa. The poly-L-lysine-coated dish method was used to prepare the specimens, and the cerium method was employed for electron-microscopic enzyme localization. Precipitates indicating enzyme activity were detected on the outer side of the external plasma membrane of the acrosomal region. This enzyme may play a role in sperm motility and male infertility.

5'-Nucleotidase↗

PubMed2000

Therapeutic cryogel removal in autoimmune disease: what is cryogel. 1982.

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Autoimmune Diseases↗

PubMed2000

Renal cell carcinoma in acquired cystic kidney disease: volume growth rate determined by helical computed tomography.

The aim of this study is to determine the growth rate and behavior of renal cell carcinoma in chronic hemodialysis patients with acquired cystic kidney disease (ACKD). Renal cell carcinomas in 17 hemodialysis patients (mean age, 52 +/- 11 years; mean hemodialysis duration, 7.2 +/- 3.3 years) with ACKD were examined with helical computed tomography (CT) for 0.5 to 6.0 years (mean, 2.1 +/- 1.9 years). The 17 renal cell carcinomas were histologically proven and graded after nephrectomy (16 patients) or autopsy (1 patient). Tumor volume was estimated by counting the number of pixels in the tumor and a 1-cm(2) area on helical computed tomographic scan using a personal computer. Estimated volume growth rates and doubling times of the carcinoma were correlated with histological grades. Fifteen of the 17 neoplasms (88%) were less than 3 cm in diameter at initial CT. The overall volume growth rate was 0.07 to 17.34 cm(3)/y (mean, 4.14 +/- 5.66 cm(3)/y), and the estimated volume-doubling time was 0. 08 to 23.31 years (mean, 5.09 +/- 6.99 years). The mean growth rate of the 3 grade 3 carcinomas was 6.01 +/- 4.50 cm(3)/y (range, 0.88 to 9.28 cm(3)/y), which was significantly greater than that of the 9 grade 1 carcinomas (0.40 +/- 0.40 cm(3)/y; range, 0.09 to 1.37 cm(3)/y) or the 5 grade 2 tumors (0.79 +/- 0.74 cm(3)/y; range, 0.12 to 2.00 cm(3)/y). Eleven of the 17 carcinomas (65%) had more than a 1-year volume-doubling time. The 3 grade 3 neoplasms and 1 of the grade 2 lesions had less than a 0.5-year doubling time.

Aged↗

PubMed2000

TI-201 imaging in a malignant peripheral nerve sheath tumor.

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Adult↗