Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “phasing”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 415 records · Page 23Linked to original sources
PubMed2002

Identification of guanfu base A hydrochloride phase I and phase II metabolites in rat bile by liquid chromatography mass spectrometry.

AIM: To study metabolites of guanfu base A hydrochloride (GFA) in rat bile. METHODS: An analytical method was developed to identify guanfu base A and its metabolites by liquid chromatography mass spectrometry (LC-MS) and electrospray ionization tandem mass spectrum (MS-MS). Rat bile was collected after iv injection of GFA. Phase I metabolite was identified by comparison with authentic standard for their retention time, molecular ion peaks, fragment ions, and UV spectrums. In order to identified Phase II conjugates, the aglycones were identified after rat bile was treated with either glucuronidase or sulfatase firstly. Phase II conjugates were also separated and determined for their molecular ions by LC-MS, at last they were verified by identifying characteristic product ions or precursor ions by MS-MS. RESULTS: Phase I metabolite, guanfu base I (GFI), was identified in rat bile. After phase II conjugates were treated with glucuronidase or sulfatase, GFA and GFI occurred in chromatograms. Quasi molecular ions m/z 606 and 510 were separated and detected in rat bile. They were indicated to be GFA glucuronide and GFA sulfate, respectively. Furthermore, GFA glucuronide was confirmed to exist in rat bile by identifying two characteristic ions, m/z 177, [M+H]+ of glucuronic acid, and m/z 430, [M+H]+ of GFA, as product ions of m/z 606. CONCLUSION: Phase I metabolite GFI and phase II conjugates, GFA glucuronide and sulfate, GFI glucuronide and sulfate, were identified in rat bile.

Alkaloids↗
PubMed1988

Plasminogen activation by tissue plasminogen activator in the presence of stimulating CNBr fragment FCB-2 of fibrinogen is a two-phase reaction. Kinetic analysis of the initial phase of slow plasmin formation.

Plasminogen activation by tissue-type plasminogen activator (t-PA) is stimulated by fibrin. In a purified system maximal fibrin-enhanced plasmin formation occurs with a delay after an initial phase of slow plasmin formation (lag phase). In the present study purified stimulating CNBr-fragment FCB-2 of fibrinogen was used, and kinetics of plasminogen activation by t-PA were analyzed with respect to the lag phase. At constant FCB-2 concentration the duration of the lag phase decreased with increasing concentrations of t-PA and plasminogen. During this period the rate of plasmin formation/min increased linearly with time with a slope dependent on the initial concentrations of FCB-2, plasminogen, and t-PA. Plasmin pretreatment of FCB-2 resulted in a dose- and time-dependent shortening of the lag phase, and at plasmin concentrations greater than or equal to 1 nM and preincubation times greater than or equal to 3 min maximal plasmin formation occurred without a lag phase. Kinetics during the phase of maximal and constant plasmin formation were not influenced by plasmin pretreatment of FCB-2. We therefore conclude that maximal t-PA-dependent plasmin formation in a system stimulated by purified FCB-2 requires plasmin modification of FCB-2.

Cyanogen Bromide↗
PubMed1988

The cooperative antioxidant role of glutathione with a lipid-soluble and a water-soluble antioxidant during peroxidation of liposomes initiated in the aqueous phase and in the lipid phase.

A study is made of the effect of GSH as a co-antioxidant with vitamin E during free radical chain autoxidation inhibition studies of dilinoleoylphosphatidylcholine (DLPC) liposomes. Oxidations are initiated in the aqueous phase with azobis(2-amidinopropane hydrochloride) and in the bilayer phase of DLPC with azobis(2,4-dimethylvaleronitrile) under known conditions of the rate of free radical chain initiation (Ri). In reactions initiated in the aqueous phase, GSH is not an efficient antioxidant when acting alone; however, in cooperation with vitamin E in the bilayers, it does effect significant extensions of the efficient induction period of vitamin E. Quantitative studies show that GSH "spares" 0.4 molecules of vitamin E in the bilayer/molecule of GSH and therefore terminates approximately 0.8 peroxyl radical chains as a co-antioxidant with vitamin E. In contrast, GSH is not an effective co-antioxidant with an efficient water-soluble antioxidant, 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylate (Trolox). GSH spares only 0.08 molecules of Trolox/molecule of GSH during autoxidation initiated in the aqueous phase with azobis(2-amidinopropane hydrochloride). The inhibition rate constant for GSH in trapping aqueous phase peroxyls is at least an order of magnitude less than that of Trolox. When peroxidation is initiated in the bilayer phase of DLPC with azobis(2,4-dimethylvaleronitrile), GSH is not an effective co-antioxidant with either vitamin E in the bilayer or Trolox in the water. Comparatively higher ratios of GSH to E (GSH/E = 50) or Trolox (GSH/Trolox = 30) are required to give significant extensions of the E or Trolox induction periods. GSH is estimated to preserve only approximately one vitamin E or Trolox molecule for a hundred GSH for peroxidations initiated in the DLPC bilayers. From the kinetic studies and GSH decay studies during inhibition periods, it is concluded that GSH does not act synergistically by regenerating ArOH from the phenoxyl, ArO, radical of vitamin E or Trolox. The mode of antioxidant action of GSH is concluded to be that of trapping peroxyl radicals in the aqueous phase and thereby indirectly sparing vitamin E in the bilayer.

Antioxidants↗
PubMed1993

Immunization of dogs with Q fever vaccines: comparison of phase I, II and phase I CMR Coxiella burnetii vaccines.

Q fever vaccines were tested in mixed breed dogs by vaccinating them with formalin-killed Coxiella burnetii whole cells (WC) in either phase I (WCI) or phase II (WCII), or the chloroform: methanol residue (CMR) subunit of phase I cells. Phase I vaccines mixed (1:1) with Freund's incomplete adjuvant (FIA) induced humoral immune responses to phases I and II antigens as measured by microagglutination assay. The CMR vaccine mixed (1:1) with FIA induced greater antigen-specific antibody levels to both phases I and II antigens than the corresponding WCI vaccine. The WCII vaccine induced antibody responses to only phase II antigens. The time course of erythema and induration after skin testing with C. burnetii antigens were suggestive of cell-mediated immunity (CMI). Although granulomas were observed with only WCI and WCII, none of the skin test antigens induced abscesses at the injection site. In contrast, axillary nodes draining the vaccine injection site developed sterile draining abscesses in all dogs by days 19 to 24 for the WCI and CMR, and day 104 for the WCII vaccines. The abscesses had resolved within 30 days after first appearance. Responses to Con A and PHA and recall antigens of lymphocytes from the blood, axillary and mesenteric nodes, and spleen at 222 days after vaccination were variable among dogs. Lymphocytes from various organs responded to one or more of the recall antigens and to both mitogens in the absence or presence of indomethacin. Although these Q fever vaccines induced humoral and CMI, either the antigens or FIA caused sterile draining abscesses.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1996

[CT findings of the irradiated intestinal wall: comparison of early vascular phase and delayed equilibrium phase].

The intestinal walls of the irradiated pelvic tumor cases of 20 patients were studied. CT manifestations of the irradiated intestinal wall were examined at the early vascular phase (examined 40 seconds after the injection of contrast medium by helical scan) and at the delayed equilibrium phase (examined 4 to 5 minutes after the injection of contrast medium by rapid sequence scan). Two characteristics were identified: intestinal wall thickening and a three-layered pattern. The former was better depicted on equilibrium phase, whereas the latter was clearer on early vascular phase. Differences in visualization of the wall thickening on vascular phase and delayed phase were statistically significant. The three-layered pattern was considered to suggest severe inflammatory edematous change of the intestinal wall. Early enhancement of the mucosal and muscular-serosal layers with less enhancement of the submucosal layer, and homogeneous distribution of the contrast medium to the interstitium on delayed phase were considered to be the reason.

Adult↗
PubMed1997

Stimulation on the positive phase of hippocampal theta rhythm induces long-term potentiation that can Be depotentiated by stimulation on the negative phase in area CA1 in vivo.

Long-term potentiation (LTP) of synaptic transmission induced by high-frequency stimulation (HFS) is considered to be a model for learning processes; however, standard HFS protocols consisting of long trains of HFS are very different from the patterns of spike firing in freely behaving animals. We have investigated the ability of brief bursts of HFS triggered at different phases of background theta rhythm to mimic more natural activity patterns. We show that a single burst of five pulses at 200 Hz given on the positive phase of tail pinch-triggered theta rhythm reliably induced LTP in the stratum radiatum of the hippocampus of urethane-anesthetized rats. Three of these bursts saturated LTP, and 10 bursts occluded the induction of LTP by long trains of HFS. Burst stimulation on the negative phase or at zero phase of theta did not induce LTP or long-term depression. In addition, stimulation with 10 bursts on the negative phase of theta reversed previously established LTP. The results show that the phase of sensory-evoked theta rhythm powerfully regulates the ability of brief HFS bursts to elicit either LTP or depotentiation of synaptic transmission. Furthermore, because complex spike activity of approximately five pulses on the positive phase of theta rhythm can be observed in freely moving rats, LTP induced by the present theta-triggered stimulation protocol might model putative synaptic plastic changes during learning more closely than standard HFS-induced LTP.

Anesthetics, Intravenous↗
PubMed1999

Evaluation of gadobenate dimeglumine in hepatocellular carcinoma: results from phase II and phase III clinical trials in Japan.

To evaluate the clinical efficacy of gadobenate dimeglumine (Gd-BOPTA)-enhanced magnetic resonance imaging for hepatocellular carcinoma (HCC), we reviewed the results of clinical phase II and III trials in Japan. Gd-BOPTA was administered at a dose of 0.1 mmol/kg to 139 patients who were suspected to have HCC. Dynamic phase images [breath-hold T1-weighted gradient echo (GRE)], spin-echo (SE) images obtained within 10 minutes of injection, and delayed breath-hold GRE images obtained 40-120 minutes after injection were evaluated. All post-contrast images were compared with T1- and T2-weighted pre-contrast images. The contrast efficacy for the dynamic study was classified as ( ) or (++) in 92.1% (128/139), in 43.1% (59/137) with SE within 10 minutes of injection, and in 43.2% (60/139) with breath-hold GRE at delayed phase. The increase in lesion-liver contrast-to-noise ratio was best at the arterial phase of dynamic breath-hold GRE. Liver signal-to-noise ratio showed a mean 52.3% increase in delayed phase. Additional information at delayed phase compared with images acquired within 10 minutes of injection (including the dynamic study) was classified as ( ) or (++) in 28.1% (39/139). With regard to safety, the overall incidence of adverse reactions was 5.0% (7/141) of the patients who were suspected to have HCC, all of whom recovered within 12 hours without any sequelae. No clinically important changes were observed in the blood and urine laboratory tests. It was concluded that Gd-BOPTA was well tolerated and effective in both dynamic study and delayed static imaging for the diagnosis of HCC.

Adult↗
PubMed1996

Okadaic acid overrides the S-phase check point and accelerates progression of G2-phase to induce premature mitosis in HeLa cells.

Okadaic acid has been shown to induce premature mitosis in HeLa cells belonging to both S-phase and G2-phase. The response is dependent on concentration and perhaps also on the differential susceptibility of cells to okadaic acid (OA). OA resembles caffeine in its ability to override the DNA-replication check point but differs in being capable of inducing premature mitosis in cells in G2-phase, thereby accelerating mitosis. Chromosomes from mitotic cells induced in G2-phase show a typical banding pattern similar to G-banding. From immunoblot experiments and survey of the relevant literature it has been argued that OA may modulate both a tyrosine kinase and a tyrosine phosphatase activity to override the S-phase checkpoint and accelerate the activation of MPF in cells in G2-phase, respectively.

Cell Cycle↗
PubMed1993

Luteal phase-characteristic induction of I3SO3-GalCer in human cervical epithelia and uterine endometria, and follicular phase-characteristic formation of a ganglioside-derived negative charge gradient in different regions of fallopian tubes.

In a series of experiments on the hormone-dependent molecular alteration in the human genital tract during the menstrual cycle, we focused our attention on a change in the negative charge due to the sulfuric acid- and sialic acid-containing glycosphingolipids. Although a ganglioside-derived negative charge was maintained in the cervical epithelia and uterine endometria at a relatively constant concentration throughout the luteal and follicular phases, I3SO3GalCer in both tissues characteristically increased in the luteal phase, indicating that the synthesis of I3SO3-GalCer in both tissues is associated with the menstrual cycle. However, I3SO3-GalCer in mucosae of the fallopian tubes in both phases was present in a concentration similar to that in the uterine endometrium in the luteal phase, and the change in the concentration did not associated with the menstrual cycle. On the other hand, although the concentrations of I3SO3-GalCer and II3NeuAc-LacCer, a major ganglioside, were similar in different regions, that is, the isthmus, ampulla and fimbriae of the fallopian tubes in the luteal phase, II3NeuAc-LacCer was present in a gradually increasing concentration from the isthmus to the fimbriae in the follicular phase, giving a gradually decreasing ratio of I3SO3GalCer to ganglioside from the uterus to the fimbriae. These findings indicate that the metabolism of sulfo- and sialoglycosphingolipids in the human genital tract is strictly controlled by estrogen and progesterone.

Cervix Uteri↗
PubMed1998

Adrenergic receptors in premenstrual dysphoric disorder: I. Platelet alpha 2 receptors: Gi protein coupling, phase of menstrual cycle, and prediction of luteal phase symptom severity.

BACKGROUND: Abnormal alpha 2-adrenergic receptor (AR) function is implicated in anxiety and depressive disorders. Premenstrual dysphoric disorder (PMDD) is characterized by anxiety and depressive symptoms, which may be associated with changes in alpha 2AR function. Previous studies on alpha 2AR function during phases of the menstrual cycle in controls and PMDD patients are inconsistent. METHODS: alpha 2AR function was examined in 16 PMDD patients and 15 controls during the follicular phase, and in 10 PMDD patients during late luteal phase. Antagonist-measured maximum binding capacity, agonist-measured receptor density in high- and low-conformational states, and agonist affinity to both states were measured. Coupling efficiency to Gi protein was estimated. RESULTS: There were no significant differences in coupling efficiency. PMDD patients had significantly low antagonist affinity; there were no differences in other binding parameters. There were no changes in alpha 2AR binding parameters between phases of menstrual cycle in PMDD women. alpha 2AR density and symptom severity were inversely related during the follicular phase in controls and patients. During luteal phase, alpha 2AR density correlated positively with symptom severity in patients. High follicular alpha 2AR density predicted more severe luteal symptoms in PMDD patients. CONCLUSIONS: These findings are discussed in view of the molecular biology of alpha 2AR, and their role in PMDD, anxiety, and depressive disorders.

Adult↗
PubMed2000

A phase I trial of cisplatin plus decitabine, a new DNA-hypomethylating agent, in patients with advanced solid tumors and a follow-up early phase II evaluation in patients with inoperable non-small cell lung cancer.

The authors describe a phase I trial of cisplatin plus decitabine, a novel DNA-hypomethylating agent, in patients with advanced solid tumors, which was followed by an early phase II evaluation of the combination in patients with inoperable non-small cell lung cancer (NSCLC). In the phase I trial, cisplatin was studied at a fixed dose of 33 mg/m2, while decitabine was escalated in four (I-IV) dose escalation levels (45, 67, 90 to 120 mg/m2, respectively) in consecutive groups of at least 3 patients per dose level. Decytabine was administered to the patients as a two-hour intravenous infusion, while cisplatin was given intravenously immediately after the end of decitabine infusion. Both agents were given on days 1-3 every 21 days. Twenty-one patients were included in the phase I trial. Dose level IV (120 mg/m2 decitabine) was considered the maximum tolerated dose (MTD), while the dose-limiting toxicities were neutropenia, thrombocytopenia and mucositis. The recommended doses for phase II trials in good- and poor-risk patients were 90 (level III) and 67 mg/m2 (level II), respectively. One short-lasting partial response was observed in a patient with cervical cancer, while two minor regression were documented in a patients with NSCLC and cervical cancer, respectively. Dose level II was selected for the phase II trial in patients with inoperable NSCLC. Fourteen consecutive patients were included in this part of the study. The median age of the patients was 57 years (range, 39-75), male/female ratio of 11/3 and a median WHO performance status 1 (0-2). The stage of disease were IIIB (5) and IV (9). Prior irradiation to the chest was given in one case. A total of 30 treatment courses were evaluable for toxicity and response, with a median of 2 courses per patient (1-4). Grade 3-4 neutropenia and thrombocytopenia were observed in about half of the cases. Mucositis, diarrhea, nausea and vomiting, and skin rash were also observed in some patients. Three minor responses were documented, which lasted for 4, 16 and 36 weeks. Median survival of patients was 15 weeks (4-38). In conclusion, the cisplatin plus decitabine combination did not exhibit significant antitumor activity in patients with NSCLC at the dose and schedule applied in this trial to justify its further evaluation in this patient population.

Adult↗
PubMed1999

A cyclin D-Cdk4 activity required for G2 phase cell cycle progression is inhibited in ultraviolet radiation-induced G2 phase delay.

Cyclin D-Cdk4 complexes have a demonstrated role in G1 phase, regulating the function of the retinoblastoma susceptibility gene product (Rb). Previously, we have shown that following treatment with low doses of UV radiation, cell lines that express wild-type p16 and Cdk4 responded with a G2 phase cell cycle delay. The UV-responsive lines contained elevated levels of p16 post-treatment, and the accumulation of p16 correlated with the G2 delay. Here we report that in UV-irradiated HeLa and A2058 cells, p16 bound Cdk4 and Cdk6 complexes with increased avidity and inhibited a cyclin D3-Cdk4 complex normally activated in late S/early G2 phase. Activation of this complex was correlated with the caffeine-induced release from the UV-induced G2 delay and a decrease in the level of p16 bound to Cdk4. Finally, overexpression of a dominant-negative mutant of Cdk4 blocked cells in G2 phase. These data indicate that the cyclin D3-Cdk4 activity is necessary for cell cycle progression through G2 phase into mitosis and that the increased binding of p16 blocks this activity and G2 phase progression after UV exposure.

Cyclin D↗
PubMed1997

Pharmacokinetic and pharmacodynamic properties of docetaxel: results of phase I and phase II trials.

The pharmacokinetics of docetaxel as investigated in Phase I and Phase II trials are discussed. Phase I trials have shown that docetaxel exhibits linear pharmacokinetics consistent with a three-compartment model. Population pharmacokinetic and pharmacodynamic analysis of the results of 22 nonrandomized Phase II trials with nonlinear mixed-effects modeling showed that the interpatient variability of docetaxel pharmacokinetics is mainly related to body surface area and hepatic function. The effect of body surface area on clearance does not present a problem, since the dose of docetaxel is adjusted for body size. However, patients with impaired liver function are at increased risk of serious adverse effects (febrile neutropenia, severe infections, severe stomatitis, and toxic death) during treatment with docetaxel 100 mg/ m2. Patients with impaired liver function should receive a reduced dose (75 mg/m2). The presence of liver metastases without impaired liver function had no effect on docetaxel's clearance or safety, suggesting that a dose of 100 mg/m2 is well tolerated in such patients. Docetaxel pharmacokinetics and pharmacodynamics determined in Phase I and Phase II trials indicate a need to adjust the dosage for body surface area and liver function.

Antineoplastic Agents, Phytogenic↗
PubMed2004

Blood to the cornual area of the uterus is mainly supplied from the ovarian artery in the follicular phase and from the uterine artery in the luteal phase.

BACKGROUND: The blood supply to the uterus is provided by the uterine and ovarian arteries, which form anastomoses. Yet the flow direction through this anastomoses and the primary source of blood supply to the tubes and uterine cornua remains unknown. To clarify this issue, we studied the spatial propagation of temperature changes following cooling of the upper vaginal area. METHODS: A thermocatheter with eight measurement points at 5-mm intervals was inserted into the uterus of nine women in the follicular phase and 11 in the luteal phase. The distal tip was positioned in the cornual area and temperatures were registered every 2 s. The vagina was then cooled for 7 min with 25 degrees C saline. RESULTS: The pattern of uterine cooling based on local counter-current transfer differed between the follicular and luteal phase. Cooling of the cornual area was significantly lower in the luteal phase compared with the follicular phase, indicating a shift in the prevailing source of arterial supply in that area following ovulation. CONCLUSIONS: The divide between the territories irrigated by the uterine and ovarian arteries moves between the follicular and luteal phase. This constitutes the first description of a functionally determined shift in the territorial divide of two vascular systems, and has numerous practical implications.

Adult↗
PubMed2004

Phase I and phase II safety and efficacy trial of intercellular adhesion molecule-1 antisense oligodeoxynucleotide (ISIS 2302) for the prevention of acute allograft rejection.

BACKGROUND: ISIS 2302, an antisense oligonucleotide that inhibits the expression of human intercellular adhesion molecule (ICAM)-1, was evaluated in combination with a cyclosporine (CsA)-prednisone (Pred) regimen first in a phase I safety and pharmacokinetic study and then in a phase II assessment of prophylaxis of acute rejection episodes in deceased donor renal allografts. METHODS: Both phase I and phase II trials were double-blinded and placebo-controlled, including 17 stable and 39 de novo patients, respectively, in time-lagged, ascending-dose regimens. Each study compared the outcomes of 8 alternate-day intravenous infusions of four ISIS 2302 dose levels (0.05, 0.5, 1.0, or 2.0 mg/kg) versus placebo (3:1 ratio). Patients were followed for 34 days (phase I) or 6 months (phase II). All transplant patients were followed for 3 years. RESULTS: ISIS 2302 produced no evident toxicity; a significant, dose-related increase in activated partial thromboplastin time was accompanied by a trend toward a decreased platelet count. ISIS 2302 did not alter the pharmacokinetic behavior of CsA. At 6 months, the rates of acute rejection episodes were 38.1% in the ISIS 2302 group versus 20.0% in the placebo group. Three-year graft survivals were similar. The mean creatinine values at 1, 2, and 3 years for all ISIS dose groups combined versus placebo over 3 years showed no significant differences. CONCLUSIONS: ISIS 2302 did not evoke side-effects and produced slightly improved renal function. However, in this pilot study, it did not further reduce the rate of acute rejection episodes or increase graft survival compared to a concentration-controlled CsA-Pred regimen.

Creatinine↗
PubMed1997

A-myb is expressed in bovine vascular smooth muscle cells during the late G1-to-S phase transition and cooperates with c-myc to mediate progression to S phase.

The Myb family of transcription factors is defined by homology within the DNA binding domain and includes c-Myb, A-Myb, and B-Myb. The protein products of the myb genes all bind the Myb-binding site (MBS) [YG(A/G)C(A/C/G)GTT(G/A)]. A-myb has been found to display a limited pattern of expression. Here we report that bovine aortic smooth muscle cells (SMCs) express A-myb. Sequence analysis of isolated bovine A-myb cDNA clones spanning the entire coding region indicated extensive homology with the human gene, including the putative transactivation domain. Expression of A-myb was cell cycle dependent; levels of A-myb RNA increased in the late G1-to-S phase transition following serum stimulation of serum-deprived quiescent SMC cultures and peaked in S phase. Nuclear run-on analysis revealed that an increased rate of transcription can account for most of the increase in A-myb RNA levels. Treatment of SMC cultures with 5,6-dichlorobenzimidazole riboside, a selective inhibitor of RNA polymerase II, indicated an approximate 4-h half-life for A-myb mRNA during the S phase of the cell cycle. Expression of A-myb by SMCs was stimulated by basic fibroblast growth factor, in a cell density-dependent fashion. Cotransfection of a human A-myb expression vector activated a multimerized MBS element-driven reporter construct approximately 30-fold in SMCs. The activity of c-myb and c-myc promoters, which both contain multiple MBS elements, were similarly transactivated, approximately 30- and 50-fold, respectively, upon cotransfection with human A-myb. Lastly, A-myb RNA levels could be increased by a combination of phorbol ester plus insulin-like growth factor 1. To test the role of myb family members in progression through the cell cycle, we comicroinjected c-myc and myb expression vectors into serum-deprived quiescent SMCs. The combination of c-myc and either A-myb or c-myb but not B-myb synergistically led to entry into S phase, whereas microinjection of any vector alone had little effect on S phase entry. Thus, these results suggest that A-myb is a potent transactivator in bovine SMCs and that its expression induces progression into S phase of the cell cycle.

Animals↗
PubMed1997

Ras activity late in G1 phase required for p27kip1 downregulation, passage through the restriction point, and entry into S phase in growth factor-stimulated NIH 3T3 fibroblasts.

It is well documented that Ras functions as a molecular switch for reentry into the cell cycle at the border between G0 and G1 by transducing extracellular growth stimuli into early G1 mitogenic signals. In the present study, we investigated the role of Ras during the late stage of the G1 phase by using NIH 3T3 (M17) fibroblasts in which the expression of a dominant negative Ras mutant, p21(Ha-Ras[Asn17]), is induced in response to dexamethasone treatment. We found that delaying the expression of Ras(Asn17) until late in the G1 phase by introducing dexamethasone 3 h after the addition of epidermal growth factor (EGF) abolished the downregulation of the p27kip1 cyclin-dependent kinase (CDK) inhibitor which normally occurred during this period, with resultant suppression of cyclin Ds/CDK4 and cyclin E/CDK2 and G1 arrest. The immunodepletion of p27kip1 completely eliminated the CDK inhibitor activity from EGF-stimulated, dexamethasone-treated cell lysate. The failure of p27kip1 downregulation and G1 arrest was also observed in cells in which Ras(Asn17) was induced after growth stimulation with a phorbol ester or alpha-thrombin and was mimicked by the addition late in the G1 phase of inhibitors for phosphatidylinositol-3-kinase. Ras-mediated downregulation of p27kip1 involved both the suppression of synthesis and the stimulation of the degradation of the protein. Unlike the earlier expression of Ras(Asn17) at the border between G0 and G1, its delayed expression did not compromise the EGF-stimulated transient activation of extracellular signal-regulated kinases or inhibit the stimulated expression of a principal D-type cyclin, cyclin D1, until close to the border between G1 and S. We conclude that Ras plays temporally distinct, phase-specific roles throughout the G1 phase and that Ras function late in G1 is required for p27kip1 downregulation and passage through the restriction point, a prerequisite for entry into the S phase.

3T3 Cells↗
PubMed1987

Longitudinal evaluation of the luteal phase and its transition into the follicular phase.

The precise patterns of LH, FSH, and PRL secretion and their correlation with estradiol (E2) and progesterone (P) secretion during the entire luteal phase have not been elucidated. To analyze in detail the secretory patterns of these hormones we performed 29 consecutive studies in 5 healthy, regularly menstruating women throughout their luteal phase [days 0 (ovulation), 2, 6, 10, and 14] and subsequent early follicular phase (day 2F). During each study plasma LH, FSH, PRL, E2, and P were measured at 10-min intervals for 6 h. Both plasma LH concentrations and LH pulse frequency declined from days 0 to 10 and increased thereafter, whereas LH pulse amplitude continued to decline throughout the luteal and early follicular phases. Plasma FSH concentrations followed a pattern similar to that of LH; however, there was a larger increase in the FSH level on days 14 and 2F. Plasma PRL levels declined initially on day 2 and again on day 14. Regression analysis indicated a positive correlation between LH concentrations and LH pulse frequency (r = 0.715; P less than 0.001) and between PRL and E2 concentrations (r = 0.528; P less than 0.01). A negative correlation was found between plasma P concentrations and both LH concentrations (r = -0.521; P less than 0.01) and LH pulse frequency (r = -0.633; P less than 0.001) and between plasma E2 and FSH concentrations (r = -0.762; P less than 0.001). Thirty-six (65%) PRL pulses and only 11 (39%) FSH pulses coincided with LH pulses. There was no clear pulsatile pattern of secretion of either E2 or P. We conclude that 1) the plasma LH, FSH, PRL, E2, and P concentrations vary markedly throughout the luteal phase; 2) the plasma LH level is largely dependent on the frequency of LH pulses; 3) plasma P decreases plasma LH by reducing the frequency of LH pulses; 4) the remarkable synchrony between PRL pulses and LH pulses implies that their secretion may be regulated by a common neuroendocrine mechanism; and 5) the preferential increase in FSH during the late luteal phase may play an important role in follicular recruitment for the subsequent cycle.

Adult↗