Biomedical subjects

W I Li

Publications and source records attributed to W I Li.

11 recordsLinked to original sources

PubMed1992

Porcine endometrial epithelial cells immortalized by transfection with origin-defective, temperature-sensitive simian virus 40 DNA.

The purpose of this study was to immortalize porcine endometrial cells and to characterize the transformed cells. Primary porcine endometrial cells were transfected with the plasmid vector (pmk16) containing SV40 DNA using a liposome-mediated method. The viral DNA was from a replication-defective, origin-minus, temperature-sensitive mutant strain (A58). One clone, designated PE-1, has been propagated for over 120 passages. PE-1 cells grown at 33C (33C cells) exhibit spindle-shaped morphology; when cultured at 40C (40C cells), they took on a polygonal or spherical shape. Morphology of 40C cells returned to the spindle shape after culture flasks were shifted back to 33C. During a 2-week period, 33C cells propagated approximately 30-fold faster than 40C cells, whereas protein concentration was higher in 40C cells. Southern blot analysis of PE-1 cells demonstrated successful integration of the ts-SV40 DNA sequence into the porcine endometrial cells, possibly at multiple sites. The presence of cytokeratin on PE-1 cell membranes was shown by immunocytochemical studies, suggesting that the PE-1 cell clone was of epithelial origin. Reverse phase (RP)-HPLC analysis of PE-1 cell extract indicated that the majority of immunoreactive beta-endorphin (ir-BEND) eluted with a hydrophobicity similar to that of synthetic BEND and alpha-N-acetylated BEND (Nac-BEND). These results demonstrate that a porcine endometrial cell line has been established, and that this cell line possesses characteristics of temperature sensitivity in cell morphology, growth rate, and protein synthesis.

Animals↗

PubMed1992

Methionine-enkephalin in a porcine endometrial cell line and its responsiveness to potassium depolarization.

Immunoreactive methionine-enkephalin (ir-MENK) has been identified in the porcine uterine fluid and endometrium. Previously, we have established a porcine endometrial cell line of epithelial origin (PE-1) by transfecting primary endometrial cells with temperature sensitive SV40 DNA. The current study was conducted to identify and characterize ir-MENK present in PE-1 cells, and to investigate the effect of KCl depolarization on the kinetics of ir-MENK secretion. PE-1 cells were cultured at 33C until confluency was reached (33C cells), after which they were incubated at 40C for 2 days (40C cells). Ir-MENK in PE-1 cells was analyzed by Sephadex G-15 gel filtration and reverse phase (RP)-HPLC. Analysis of 40C cell extract by Sephadex G-15 and RP-HPLC indicated that the major portion of ir-MENK present in PE-1 cells was eluted at a position similar to that of synthetic MENK. The effect of temperature on ir-MENK synthesis in PE-1 cells was examined by measuring ir-MENK content in 33C and 40C cells over a 14-day culture period. Compared to 33C cells, 40C cells maintained higher and steadier levels of ir-MENK, suggesting that synthesis of ir-MENK is temperature sensitive. KCl stimulated ir-MENK secretion at all concentrations tested (5-60 mM for 60 min), with 30 mM being the optimal concentration. Temporal analysis of ir-MENK secretion showed that incubation for 60 min with 30 mM KCl allowed maximal secretion. Secretion of ir-MENK from PE-1 cells resulted in depletion of ir-MENK in cell content. These results demonstrate that PE-1 cells contain ir-MENK which is biochemically similar to synthetic MENK, PE-1 cells synthesize ir-MENK in a temperature sensitive manner, and these cells secrete ir-MENK upon KCl stimulation.

Animals↗

PubMed1992

Endometrial immunoreactive beta-endorphin increases during mid-estrous cycle and early pregnancy in gilts.

Immunoreactive (ir) beta-endorphin (BEND) was recently identified in porcine uterine fluids. In the study reported here, we examined the hypothesis that porcine endometrium serves as a source of uterine fluid ir-BEND during the estrous cycle and early pregnancy. Endometrial ir-BEND was chromatographically characterized, sites of ir-BEND synthesis were immunocytochemically localized, and concentrations of endometrial ir-BEND during the estrous cycle and early pregnancy were measured. Sephadex G-50 chromatographic profiles of endometrial extracts from Day 15 of the estrous cycle revealed three distinct peaks of ir-BEND, with the first peak occurring near void volume and the second and third peaks coinciding with standard porcine beta-lipotropin and standard porcine BEND, respectively. Reverse-phase HPLC C18 chromatographic profiles indicated that endometrial ir-BEND contained both standard BEND and alpha-N-acetylated BEND. Immunocytochemical studies demonstrated ir-BEND in the surface and glandular epithelial cells of the endometrium, with immunostaining most prominent in the apical portion of epithelial cells. Concentrations of ir-BEND in endometrial tissues were higher on Days 14-15 than on Days 8-12 during the estrous cycle and pregnancy (p less than 0.05); however, values were not different in pregnant and cyclic gilts. Biochemical and immunocytochemical evidence supports our hypothesis that ir-BEND present in uterine fluids is derived from the endometrium. The increase in endometrial ir-BEND concentration during Days 14-15 in cyclic and pregnant gilts indicates that ovarian steroids may influence the synthesis of endometrial ir-BEND.

Animals↗

PubMed1991

In vitro studies of deformation and adhesion properties of transformed cells.

The micropipet aspiration technique and the parallel-plate flow chamber were used to investigate the deformation and detachment properties, respectively, of normal and transformed rat fibroblasts. The normal Cloned Rat Embryo Fibroblasts (CREF) cell line was transfected with the T24 ras oncogene to produce the transformed cell line CREF T24. The CREF T24 cell line was transfected with a Kirsten ras revertant gene (K-rev 1a suppressor) to produce the CT24HKB1 cells, which have the same morphological characteristics as the cells in the CREF line. The cells utilized in this investigation were derived from the parent cell line CREF, the only differences being the presence or absence of the T24 ras oncogene and the Kirsten ras revertant gene. The detachment and deformation properties, therefore, could be related to the metastatic phenotype of the cell rather than inherent differences between disparate cell lines. Results indicated that transfecting the CREF cell line with the ras oncogene greatly modified the detachment and deformation properties. The CREF T24 cells were more easily detached from normal cells and were 50% more deformable. Both CREF and CT24HKB1 showed similar detachment properties. Based on these results, it is speculated that K-rev 1a reversed ras-induced membrane alterations in these cells. Preliminary investigations have demonstrated that both CREF and CREF T24 cells in different phases of the cell cycle differed in morphological characteristics. However, the majority of the cells within a given cell line showed similar deformation characteristics. Current investigations are focusing on characterization of both detachment and deformation properties of these cells as a function of the cell cycle using synchronization techniques.

Animals↗

PubMed1991

Spectrophotometric prediction of the dissolution rate of carbamazepine tablets.

A near-infrared (IR) spectrophotometer, integrating optics, and parallel-vector supercomputer are employed to develop a mathematical model that predicts the dissolution rate of individual intact tablets from near-IR spectra (r2 = 0.985). Each tablet can be analyzed nondestructively by the spectrophotometer in less than 1 min. The model permits hundreds of near-IR wavelengths to be used in the determination of dissolution rate, leading to increased accuracy.

Carbamazepine↗

PubMed1991

Synthesis and secretion of immunoreactive methionine-enkephalin from rabbit reproductive tissues in vivo and in vitro.

The expression of the proenkephalin gene has been demonstrated in the reproductive tissues of several animal species. The objectives of the experiments reported here were to (a) examine the presence of immunoreactive methionine-enkephalin (ir-MENK) in rabbit ovary, oviduct, and uterus and in a rabbit endometrial cell line (HRE-H9), (b) characterize ir-MENK biochemically, (c) investigate the effect of eCG + hCG treatment on the synthesis and secretion of ir-MENK in vivo, and (d) study the effect of K+ depolarization on the secretion of ir-MENK from HRE-H9 cells. Uterine fluid was collected by flushing the uterine lumen with saline. Reproductive tissues and HRE-H9 cells were extracted with 0.1 N acetic acid. Both the uterine fluid and extracts of uterus, ovary, oviduct, and HRE-H9 cells exhibited inhibition curves parallel to that of authentic MENK in the MENK RIA system. Sephadex G-15 gel filtration profiles indicated that in the extracts of rabbit uterus and HRE-H9 cells, most ir-MENK co-eluted with standard MENK, with a minor portion eluting near the void volume (Vo). Reverse-phase-HPLC (RP-HPLC) profiles showed a major peak coinciding with standard MENK, plus a minor peak of highly hydrophilic ir-MENK. The effect of eCG + hCG treatment was studied by i.m. injection of eCG (150 IU), followed by i.v. injection of hCG (75 IU) 4 days later. Ir-MENK concentration in the uteri and ovaries was significantly (p less than 0.05) increased (9.06 +/- 1.89 and 2.05 +/- 0.32 ng/mg protein, respectively), compared to control levels (2.31 +/- 0.86 and 0.24 +/- 0.77).(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1991

Immunoreactive methionine-enkephalin secretion by porcine uterus.

The current study examined the presence of immunoreactive methionine-enkephalin (ir-MENK) in porcine uterine fluid and endometrial extracts, characterized ir-MENK biochemically, and investigated the effect of ovarian steroids on uterine secretion of ir-MENK. Porcine uterine fluid was collected by flushing the uterine lumen with saline. Endometrial tissues were extracted with acetic acid. Both uterine fluid and endometrial extracts exhibited inhibition curves parallel to that of authentic MENK in the MENK RIA system. Sephadex G-15 gel filtration chromatographic profiles indicated that both concentrated uterine fluid and endometrial extracts contained two peaks of ir-MENK, a major peak which coeluted with standard MENK, and a minor peak eluting near the void volume (Vo). Reverse phase-HPLC chromatographic profiles also demonstrated two peaks of ir-MENK for concentrated uterine fluid and endometrial extracts, a major peak which coincided with standard MENK, plus a highly hydrophilic peak. The effect of ovarian steroids on the uterine secretion of ir-MENK was examined by measuring ir-MENK in uterine fluids from cyclic and pregnant gilts as well as ovariectomized, ovarian steroid-treated gilts. Day effects (P less than 0.01) were detected for cyclic and pregnant gilts, since values for ir-MENK increased between days 8 and 14 after onset of estrus. In ovariectomized gilts, treatment with progesterone (P4) increased the uterine secretion of ir-MENK (202 +/- 9 vs. 65 +/- 4 pg/ml for control, P less than 0.05). The combined treatment of P4 and estradiol did not further enhance secretion of ir-MENK, while treatment with estradiol did not alter ir-MENK levels relative to values for control gilts. These results indicate the presence of ir-MENK in porcine uterine fluid and endometrium, and suggest that uterine secretion of ir-MENK is regulated primarily by P4.

Animals↗

PubMed1991

Viscoelastic properties of transformed cells: role in tumor cell progression and metastasis formation.

The micropipette aspiration technique was used to investigate the deformation properties of a panel of nontransformed and transformed rat fibroblasts derived from the same normal cell line. In this method, a step negative pressure is applied to the cell via a micropipette and the aspiration distance into the pipette as a function of time is determined using video techniques. A standard solid viscoelastic model was then used to analyze the viscoelastic properties of the cell. From these results, it is concluded that a direct correlation exists between an increase in deformability and progression of the transformed phenotype from a nontumorigenic cell line into a tumorigenic, metastatic cell line.

Animals↗

PubMed1989

Characterization of a temperature-sensitive beta-endorphin-secreting transformed endometrial cell line.

An endometrial cell line (HRE-H9) was established and characterized to study the mechanism by which gene expression of POMC-derived peptides is controlled in uterine tissues. The HRE-H9 cell line was isolated by transforming primary rabbit endometrial cell cultures, derived from hCG-treated pseudopregnant rabbits, with a temperature-sensitive A209 mutant (tsA209) simian virus 40 at a permissive temperature (33 C). The resulting cells exhibited temperature sensitivity in growth and synthesis of immunoreactive beta-endorphin (ir-beta END). The ir-beta-END present in the cell extracts and culture media was assayed by a specific beta END RIA. Sephadex G-50 gel filtration chromatography of the transformed cell extracts showed three peaks of beta END immunoreactivity. The first peak eluted at the void volume, the second peak coeluted with the beta-lipotropin standard, and the third peak coincided with the porcine beta END standard, ir-beta-END was also detectable in endometrial culture media, suggesting that the transformed endometrial cells secreted POMC-derived peptides. Our data indicate that the tsA209 mutant virus-transformed endometrial cell line provides a suitable model for study of the synthesis and regulation of POMC-derived peptides in extrapituitary tissues.

Animals↗

PubMed1979

Reversal of exertional hypotension after coronary bypass grafting.

A decrease in systolic blood pressure that occurs with treadmill exercise testing may be a sign of reversible ischemic left ventricular dysfunction. To test this hypothesis, we examined retrospectively the postoperative treadmill responses of 37 patients who had exertional hypotension (end exercise systolic blood pressure less than or equal to initial preexercise levels) before coronary arterial bypass grafting. This group of 37 patients was characterized preoperatively by an abnormal exercise electrocardiogram (36 patients), multiple vessel occlusive disease (36 patients) and a normal ejection fraction at rest (32 patients). Postoperative exercise tests showed improvement in hemodynamic and electrocardiographic changes with reversal of exertional hypotension (33 patients), and conversion to a normal exercise electrocardiogram (29 patients). Coronary bypass surgery can be expected to reverse exertional hypotension in patients with symptomatic angina pectoris and evidence of ischemia in the exercise electrocardiogram.

Adult↗

PubMed1979

Intraoperative management of renal cell carcinoma with supradiaphragmatic caval extension.

A new technique for inferior vena caval tumor thrombectomy is described. Vascular isolation of the cava from the right atrium to the pelvis is achieved by temporary circulatory arrest of the lower torso. Removal of the neoplastic thrombus under direct vision with minimal blood loss was accomplished in a patient with renal carcinoma, whose tumor extended into the intrapericardial vena cava.

Adenocarcinoma↗