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Biomedical subjects

Y Hu

Publications and source records attributed to Y Hu.

At least 361 records · Page 20Linked to original sources
PubMed1998

Maximizing pregnancy rates and limiting higher-order multiple conceptions by determining the optimal number of embryos to transfer based on quality.

OBJECTIVE: To define statistical thresholds for the number of embryos to be transferred to achieve an optimal pregnancy rate and keep higher-order multiple conceptions (pregnancy with more than two fetal sacs with cardiac activity) within an acceptable limit. DESIGN: A retrospective review of patient records. SETTING: Private practice assisted reproductive technology (ART) facility. PATIENT(S): Seven hundred fifty-four consecutive patients who underwent IVF-ET from 1994-1996. INTERVENTION(S): Embryo grading and score system used on day 3 of embryo transfer. MAIN OUTCOME MEASURE(S): Implantation, pregnancy, and multiple conception rates. RESULT(S): For women < or =35 years old, transfer of up to four poor-quality, two fair-quality, or two good-quality embryos is optimal to eliminate any risk of higher-order multiple pregnancies. Transfer of four poor-quality, three fair-quality, or two good-quality embryos is recommended for women 36 to 39 years old. In women who are > or =40 years old, five embryos need to be transferred regardless of embryo quality. CONCLUSION(S): The mean cumulative embryo score can be used as a reference to determine an optimal number of embryos to transfer and to predict pregnancy outcome.

Adult↗
PubMed1998

Timed analysis of the nuclear maturation of oocytes in early preantral mouse follicle culture supplemented with recombinant gonadotropin.

OBJECTIVE: To analyze the effects of combined FSH and variable doses of LH on the nuclear maturity and capacity to resume meiosis in oocytes from preantral follicles from prepubertal mice. DESIGN: Prospective, randomized, and controlled in vitro laboratory experiment. SETTING: Academic research environment. INTERVENTION(S): Meiosis was studied after somatic cell removal or after stimulation with hCG plus epidermal growth factor in three culture conditions: maturation medium with FSH alone and with two different doses of LH. MAIN OUTCOME MEASURE(S): The nuclear maturation of the oocytes and the E2, progesterone, and alpha-specific inhibin content of the conditioned medium. RESULT(S): Somatic cell removal and hormonal stimulation were equally effective in inducing germinal vesicle breakdown, but the hormonal stimulus was essential for the completion of meiosis, which was maximal (70%) on day 13 of culture. Continuous addition of LH to FSH during the oocytes' growth made them more prone to spontaneous resumption of meiosis I but resulted in a higher proportion of oocytes reaching the completion of meiosis. Estradiol and progesterone measurements demonstrated that the presence of LH influences luteinization. CONCLUSION(S): In contrast to oocytes grown in vivo, cumulus cell removal by itself is an insufficient stimulus for oocytes cultured in vitro to complete meiosis. Timed stimulation with hCG and epidermal growth factor increases nuclear maturation rates. A maximum number of metaphase II oocytes are obtained after a 13-day in vitro growth period when LH is added to the maturation medium.

Animals↗
PubMed1998

Continuous glucose monitoring in the free-moving rat.

The aim of this work was to set up an experimental model of glycemic fluctuations for assessing in the conscious freely moving rat, the performance of a continuous glucose-monitoring system, using a pocket-calculator-size electronic control unit and a miniaturized subcutaneous glucose sensor. The well-known triphasic glycemic pattern following streptozotocin injection (initial peak and secondary hypoglycemia preceding the establishment of permanent hyperglycemia) was used as a way to obtain spontaneous changes in blood glucose level over a wide concentration range. This report demonstrates that streptozotocin injection produced highly reproducible changes in the current generated by the sensor: an initial peak and a secondary nadir, during which blood sampling provided the evidence of hyperglycemia associated with immunoreactive hypoinsulinemia, and of hypoglycemia associated with hyperinsulinemia, respectively. This reproducible experimental model should be valuable for the assessment of a continuous glucose-monitoring system.

Animals↗
PubMed1998

Donor-specific stimulation of peripheral blood mononuclear cells from recipients of orthotopic liver transplants is associated, in the absence of rejection, with type-2 cytokine production.

In this study, we examined the cytokine production by human peripheral blood mononuclear cells (PBM) from recipients of orthotopic liver transplants which had been stimulated by donor-specific alloantigen. Levels of interleukin (IL)-2, IL-4, interferon (IFN)-gamma, IL-10 and transforming growth factor (TGF)-beta produced in vitro from PBM of 15 transplant recipients at 5-7 months post transplantation were analysed after donor-specific, third-party, or non-specific stimulation. Mononuclear cell proliferation in response to stimulation and cytokine mRNA from the cell cultures were assayed. Donor-specific antigen was obtained from donor spleen cells which had been obtained and frozen in liquid nitrogen at the time of organ retrieval. Third-party restimulation used equivalent numbers of spleen cells pooled from the other 14 organ donors. Cytokine production was correlated with the clinical condition of the patient, including biopsy results when available, and biochemical data. The data show a highly significant correlation between the donor-specific- and third-party- stimulated IL-4 and IL-10 production from recipient PBM with stable liver graft function as assessed by histopathology and/or biochemistry. This correlation was independent of level of immunosuppression. These data strongly support a role for IL-4 and/or IL-10 in the induction and/or maintenance of tolerance to human liver allografts. Measurement of the levels of these cytokines from recipient PBM after donor-specific antigen stimulation in vitro may be a useful test for monitoring for acute allograft rejection.

Adjuvants, Immunologic↗
PubMed1998

Abundance of platelet-derived growth factors (PDGFs), PDGF receptors and activation of mitogen-activated protein kinases in brain decline with age.

Platelet-derived growth factors (PDGFs) specifically bind to PDGF receptors (PDGFRs), resulting in their activation via autophosphorylation and subsequent triggering of a cascade of phosphorylation events that include mitogen-activated protein (MAP) kinases. Most of our knowledge concerning MAP kinase activation comes from studies of cultured cells in vitro, and little is known about their activation in vivo. In the present study, we determined PDGF and PDGFR levels and MAP kinase activities, including extracellular signal-regulated protein kinases (ERK) and c-Jun NH2-terminal protein kinases (JNK) or stress-activated protein kinases (SAPK) in brain of young and older mice. Both PDGF and PDGFR proteins were most abundant in protein extracts from brain (cerebral cortex) among tissues of heart, liver, spleen, lung and kidney, as determined by Western blot analysis. PDGFR proteins in brain differed significantly between young (1 or 8 weeks) and older (14 months) mice and PDGFR phosphorylation was seen in all age groups examined by a specific antibody against phosphotyrosine. The highest activity ERK2 was also observed in brain tissues, and this activity declined with age, although ERK1 and ERK2 protein levels were not significantly altered during development and aging. Furthermore, the activity and amount of JNK/SAPK proteins were the most abundant in brain tissues and did not change with age. Thus, our findings demonstrate that the highest levels of PDGFs and PDGFRs existed in brain, and constitutive activation of MAP kinases declined with age, suggesting that signal pathways mediated by PDGF-MAP kinase cascades are important components in coordinating growth and differentiation of neurone and glial cells during development and aging.

Aging↗
PubMed1998

Decline of shear stress-induced activation of extracellular signal-regulated kinases, but not stress-activated protein kinases, in in vitro propagated endothelial cells.

We investigated the involvement of mitogen-activated protein kinase (MAPK) signal transduction pathways in human endothelial cells in response to shear stress and alterations of these kinases in in vitro-propagated endothelial cells (ECs). Potent activation (10-fold) of extracellular signal-regulated kinase (ERK2), a member of the MAPK family, occurred within 10 min of shear stress (5 dynes/cm2), whereupon rapid inactivation ensued. Shear stress also induced activation of stress-activated protein kinase (SAPK) or c-Jun NH2-terminal protein kinase (JNK) in ECs. Suramin pretreatment completely inhibited shear stress stimulation of ERK2, but not SAPK/JNK, highlighting a role for growth factor receptors in ERK activation. Translocation of ERK2 from the cytoplasm to the nucleus was observed in shear-stressed endothelial cells. In addition, we compared activities of MAPKs in shear-stressed cells derived from passages 4 and 10 (older). The magnitude of ERK2 activation was significantly lower in aged ECs compared to those of passage 4, while SAPK/JNK was not altered in the in vitro aged ECs. A similar level of ERK2 activation was found in both young and older cells stimulated with phorbol-12-myristate-13-acetate (PMA), indicating an age-related alteration of the plasma membrane. Taken together, these findings suggest that MAP kinase activation may be crucial for the expression of many genes in ECs stimulated by shear stress, and that an alteration in MAPK activities could contribute to the age-related decline in proliferative capacity.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1998

Mutation of E2f-1 suppresses apoptosis and inappropriate S phase entry and extends survival of Rb-deficient mouse embryos.

Mice mutant for the Rb tumor suppressor gene die in mid-gestation with defects in erythropoiesis, cell cycle control, and apoptosis. We show here that embryos mutant for both Rb and its downstream target E2f-1 demonstrate significant suppression of apoptosis and S phase entry in certain tissues compared to Rb mutants, implicating E2f-1 as a critical mediator of these effects. Up-regulation of the p53 pathway, required for cell death in these cells in Rb mutants, is also suppressed in the Rb/E2f-1 double mutants. However, double mutants have defects in cell cycle regulation and apoptosis in some tissues and die at approximately E17.0 with anemia and defective skeletal muscle and lung development, demonstrating that E2F-1 regulation is not the sole function of pRB in development.

Animals↗
PubMed1998

Identification, sequence analysis and expression studies of novel anther-specific genes of Arabidopsis thaliana.

Relatively little is known about pollen development at the molecular level. For the purpose of gaining understanding of the molecular control of pollen development, a number of Arabidopsis cDNA fragments were isolated using subtractive hybridizations. DNA and RNA hybridizations and sequence analyses indicate that we have isolated cDNAs representing 13 genes. Sequences for 8 of these genes are novel, while those for the remaining 5 genes have substantial similarity to genes previously reported as anther- or pollen-specific. RNA in situ hybridizations with 5 genes revealed that four of them are tapetum-specific with differing temporal expression patterns during pollen development and one is pollen-specific within the flower. Sequence analysis of full-length cDNAs showed that one of the novel genes, ATA7, encodes a protein related to lipid transfer proteins. Another gene, ATA20, encodes a protein with novel repeat sequences and a glycine-rich domain that shares a predicted structure with a known cell wall protein. The full-length ATA27 cDNA encodes a protein similar to the BGL4 beta-glucosidase from Brassica napus. The ATA27 protein is predicted to have an ER retention signal and an acidic isoelectric point, suggesting that it may be localized to the ER lumen. This may be a means of compartmentalization from its substrate(s). Our studies demonstrate that subtractive hybridizations can be used to identify previously unknown genes, which should be valuable tools for further study of pollen and anther development and function.

Amino Acid Sequence↗
PubMed1998

Prolactin-induced expression of TATA-less cyclin D3 gene is mediated by Sp1 and AP2.

The TATA-less cyclin D3 gene is stimulated by prolactin (PRL) in rat Nb2 T-lymphocytes. DNA mobility shift assay and luciferase transfection analysis revealed that the gene has a potential promoter region, two enhancer domanins and a silencer region. The promoter region possessed three Sp1 sites and two AP2 sites, and both enhancer regions had one AP2 binding site each. Analysis using oligonucleotide probes and purified AP2 protein has suggested that the expression of rat cyclin D3 gene is promoted by Sp1 and AP2 factors.

Animals↗
PubMed1998

Co-culture with assisted hatching of human embryos using Buffalo rat liver cells.

Commercially obtained Buffalo rat liver (BRL) cells were grown in monolayer culture. The effect of BRL cell co-culture with assisted hatching on embryo development, implantation and pregnancy was investigated in a population of 200 'first-time' in-vitro fertilization (IVF) patients, subdivided into three groups according to the methods of fertilization [IVF; intracytoplasmic sperm injection (ICSI); ICSI/IVF]. Assisted hatching was performed on all embryos chosen for transfer. Following co-culture, the overall embryo quality, implantation rate and pregnancy rates were not significantly different from the controls. However, when grouped according to fertilization method, co-culture was found to have an impact on pregnancy and implantation rates in the group undergoing conventional IVF. Using co-culture with assisted hatching, we were able to achieve a 58% (38/65) clinical pregnancy rate with a 49% (32/65) live birth rate and a 26% (60/235) implantation rate. No changes in the pregnancy and implantation rates were apparent in ICSI or ICSI/IVF subgroups. This is the first prospective, randomly controlled study which reports the use of BRL cell co-culture for human IVF for a large number of patients undergoing IVF for the first time.

Adult↗
PubMed1998

Epidermal growth factor combined with recombinant human chorionic gonadotrophin improves meiotic progression in mouse follicle-enclosed oocyte culture.

Using a mouse early preantral follicle culture system, mature full grown oocytes, arrested in prophase I of meiosis, were produced after 12 days using a recombinant gonadotrophin-supplemented medium. This culture medium does not mimic the normal extracellular environment of the oocyte and might therefore modify meiotic regulation and more particularly progression to metaphase II (MII). The aim of this study was to optimize the treatment using recombinant stimulatory ligands which were known to induce germinal vesicle breakdown (GVBD) and completion of meiosis I, metaphase II (MII), namely recombinant follicle stimulating hormone (r-FSH), chorionic gonadotrophin (r-HCG) and epidermal growth factor (EGF). Full-grown intrafollicular oocytes could not resume meiosis when the 'ovulatory' stimulus was r-FSH, used at a 100 times higher dose than during culture. r-FSH did not increase progesterone production. When 1.5 IU/ml r-HCG was used as meiotic trigger, germinal vesicle breakdown was obtained in 95% of the oocytes 64% of which extruded a first polar body. r-HCG induced a dramatic increase in progesterone production. When EGF was administered as sole stimulus on day 12 to the attached follicle-enclosed oocytes, only doses > or =5 ng/ml could cause GVBD, although less effectively than r-HCG (45 versus 95%; P < 0.0001). Oocytes undergoing GVBD by the EGF pulse reached metaphase II at a rate of 54% (not significant versus r-HCG). EGF did not stimulate progesterone production. Addition of increasing doses of EGF (0.5; 5; 10; 50 ng/ml) to r-HCG did not increase the GVBD-rate, but EGF doses >5 ng/ml improved MI to MII transition (P=0.027), thereby improving the final yield of MII oocytes by 12.5%. These data show that up to a dose of 50 ng/ml, EGF on its own could only override the somatic inhibitory stimuli in less than half of the cultured follicles. However, in addition to HCG, EGF (25 ng/ml) had a stimulatory effect on completing the first meiotic division. It was concluded that, under the present culture conditions, EGF in combination with HCG provided optimal nuclear maturation.

Animals↗
PubMed1998

Recombinant luteinizing hormone as a survival and differentiation factor increases oocyte maturation in recombinant follicle stimulating hormone-supplemented mouse preantral follicle culture.

The effects of recombinant luteinizing hormone (rLH) and recombinant follicle stimulating hormone (rFSH) both separately and together were analysed on the in-vitro maturation of a well-defined class of mouse early preantral follicles. Metaphase-II (MII) oocytes were only obtained when rLH or rFSH or a combination of both hormones was added to a standard rich culture medium at constant doses throughout the culture period. Without gonadotrophins only 8% of the follicles reached day 12 of culture. Follicle survival was optimal (between 90 and 100%) when rFSH was used; significantly fewer follicles were kept intact when rLH was added as the only supplement (54%; P < 0.001). Theca cells played a fundamental role in follicle survival when rLH was the only supplement. Presence of rLH significantly influenced the formation of antral-like cavities: on day 12, 59% of follicles cultured with rLH + rFSH had a large cavity, as compared with only 21% in cultures with rFSH only (P < 0.0001). The rHCG stimulated germinal vesicle breakdown rate from the different gonadotrophin-supplemented cultures on day 12 was not significantly different (FSH: 89%; LH: 70%; FSH/LH: 83%). However, addition of rLH to rFSH did significantly improve (P < 0.05) the completion of the first meiotic division up to the MII stage (FSH: 46%; LH: 39%; FSH/LH: 76%). Interfollicle differences in steroid production were very large and depended on the presence of theca cells and on the kind of gonadotrophin supplemented. Without theca cells, oestrogen concentrations in rFSH-supplemented cultures were 100 times lower than their theca-containing counterparts during the entire culture period. From culture day 9 onwards most conditioned media had increased basal progesterone concentrations. This study points up the in-vitro role of LH on the differentiation of follicle-like structures and the oocyte's meiotic maturation and suggests an important role for theca cells in providing modulators of in-vitro gonadotrophin action.

Animals↗
PubMed1998

Activation of PDGF receptor alpha in vascular smooth muscle cells by mechanical stress.

Hypertension increases mechanical force on the arterial wall by as much as 30%, resulting in marked alterations in signal transductions and gene expression in vascular smooth muscle cells (VSMCs) that contribute to matrix protein synthesis, cell proliferation, and differentiation. How the mechanical stimuli are converted into a biological signal in cells has yet to be studied. We investigated the role of both cyclic strain and shear stresses in initiating the cellular signaling on cultured VSMCs and found that mechanical forces evoked activation of mitogen-activated protein kinases, followed by enhanced DNA binding activity of transcription factor AP-1. Physical forces rapidly induced phosphorylation of platelet-derived growth factor receptor (PDGFR) alpha, an activated state. When GRB2, an adapter protein, was immunoprecipitated from treated VSMCs followed by Western blot analysis with anti-phosphotyrosine, -PDGFR alpha, and -GRB2 antibodies, respectively, phosphotyrosine positive staining was observed on PDGFR alpha bands of the same blot in stretch-stressed VSMCs, supporting the mechanical stress-induced activation of PDGFR alpha. Conditioned medium from stretch-stressed VSMCs did not result in PDGFR alpha phosphorylation, and antibodies binding to all forms of PDGFs did not block stress-induced PDGFR alpha activation. Thus, mechanical stresses may directly perturb the cell surface or alter receptor conformation, thereby initiating signaling pathways normally used by growth factors.

Adaptor Proteins, Signal Transducing↗
PubMed1998

Expression and distribution of heme oxygenase-2 mRNA and protein in rat kidney.

Recent studies suggest that carbon monoxide (CO), which is formed by the enzyme heme oxygenase (HO) during the conversion of heme to biliverdin, shares some of the chemical and biological properties of nitric oxide (NO) and may play roles similar to those of NO. Heme oxygenase activity in the kidney has been reported for many years, and there are some reports on the expression of mRNA for two HO isozymes (HO-1 and HO-2) and cellular localization of HO-1 protein. However, cellular localization of HO-2 protein in the kidney under normal conditions has not been reported. In the present study we examined the expression and distribution of HO-2 mRNA and HO-2 protein in rat kidney using RNA protection assay and light and electron immunocytochemistry. RNA protection assay confirmed constitutive expression of HO-2 transcript in rat kidney. HO-2 immunoreactivity was selectively found in epithelial cells of the thick ascending limb and distal convoluted tubule, connecting tubule cells, and principal cells of the collecting duct. These results suggest that HO-2 is synthesized in the kidney and that HO-2 in the epithelial cells of renal tubules may serve as a source for CO generation under normal conditions.

Animals↗
PubMed1998

Use of long RT-PCR to characterize splice variant mRNAs.

Recent advances in long reverse transcription (RT)-PCR technology allow the copying of full-length coding regions of large mRNAs in one step. Using long RT-PCR, one can be certain that a given cDNA is derived from a single mRNA. In what to our knowledge is a novel application, we can isolate and characterize splice variants for any given mRNA in a systematic manner. We optimized long RT-PCR to copy the full-length coding region of human multidrug resistance (MDR1) mRNA or the major vault protein (MVP) mRNA in one step, so that only one full-length PCR product was synthesized in each case. Such stringent conditions are necessary to ensure that smaller than full-length products derived from total cell RNA are true splice variants. Twenty MDR1 double-stranded (ds) cDNAs, isolated from either the full-length or one prominent splice-variant DNA band, visualized on agarose gels, were cloned and sequenced. Two were full-length, wild-type in sequence as expected, and the rest were splice-variant mRNAs. Fourteen of the clones were identical and encoded a prominent splice-variant mRNA that can be detected in two tumor cell lines. This approach is shown to be generally applicable to the systematic analysis of splice-variant mRNAs derived from any gene.

Alternative Splicing↗
PubMed1998

Signal transduction in arteriosclerosis: mechanical stress-activated MAP kinases in vascular smooth muscle cells (review).

Vascular smooth muscle cell (SMC) proliferation is a key event in the development of (spontaneous) atherosclerosis, hypertension-related arteriosclerosis, angioplasty-induced restenosis and venous bypass graft arteriosclerosis. Many factors or environmental stimuli are believed to be responsible for SMC growth or hypertrophy in the vessel wall. How these environmental stimuli or signals applied onto the surface of SMCs are transduced into the cell nucleus resulting in quantitative and qualitative changes in gene expression in SMCs of arterial walls is largely unknown. Mitogen-activated protein (MAP) kinases are rapidly activated in cells stimulated with various extracellular signals by dual phosphorylation of tyrosine and threonine residues. They are thought to play a pivotal role in transmitting transmembrane signals required for cell growth and differentiation. Recent studies have focused on the signalling events in vascular tissues in vivo and in cultured SMCs in vitro. It has been demonstrated that acute hypertension and angioplasty rapidly induced MAP kinase activation in the arterial wall. Kinase activation is followed by an increase in c-fos and c-jun gene expression and enhanced transcription factor AP-1 DNA-binding activity. A similar MAP kinase activation can be mimicked in in vitro cultured SMCs stimulated by either shear stress or cyclic strain stretch, suggesting direct effects of mechanical force. Interestingly, physical forces rapidly resulted in phosphorylation of platelet-derived growth factor (PDGF) receptor, an activated state, in cultured SMCs. Thus, mechanical stresses may directly perturb the cell surface or alter receptor conformation, thereby initiating signalling pathways usually used by growth factors. These findings have significantly enhanced our knowledge concerning the pathogenesis of arteriosclerosis and provide a basis for therapeutic intervention on vascular diseases.

Angioplasty↗
PubMed1998

[The expressions of p16, CDK4 and PCNA proteins in trophoblastic tumors].

OBJECTIVE: To evaluate the relationship between the regulatory factors in G1 phase of cell cycle and the cacrinogenesis of the trophoblastic cells. METHODS: The expressions of p16, cyclin-dependent kinase 4 (CDK4), proliferation cell nuclear antigen (PCNA) proteins in 18 cases of trophoblastic tumors, 30 cases of hydatidiform mole and 30 cases of normal villi were studied by immunohistochemical methods. RESULTS: The expressions of p16 protein between malignant trophoblastic tumors and normal villi were significantly different (P < 0.05). PCNA positive rate in p16 positive samples were markedly lower than that in negative samples (P < 0.05). The expressions of CDK4 among normal villi, hydatiform mole and trophoblastic tumors were not significantly different. Patients with positive p16 expression had a higher 3 year survival rate. CONCLUSION: p16 protein may inhibit trophoblastic cells proliferation, and p16 gene mutation may be an important factor in carcinogenesis of trophoblastic cells and proliferation out of control. P16 protein expression examination is helpful in prediction of patients prognosis.

Choriocarcinoma↗