PubMed Health⌕ Search

Biomedical subjects

G S Schultz

Publications and source records attributed to G S Schultz.

88 records · Page 5Linked to original sources
PubMed1984

Intrauterine proximity to male fetuses predetermines level of epidermal growth factor in submandibular glands of adult female mice.

Morphological, behavioral, and physiological masculinization of adult female mice that developed in utero between two male fetuses (2M females) has been previously attributed to the significantly higher concentration of testosterone in their fetal blood and amniotic fluid than that in female mice which had not been contiguous to males in utero (0M females). Serum testosterone levels of adult 2M and 0M females are not significantly different. To determine whether exposure of fetuses to different levels of testosterone during prenatal development alters adult biochemical parameters of a system responsive to testosterone, the level of epidermal growth factor (EGF) was measured by radioreceptor assay in the submandibular glands of adult CF-1 mice of known intrauterine position. The concentration of EGF was significantly higher (P less than 0.05) in the glands of 2M females (mean +/- SEM, 0.36 +/- 0.14 nmol/mg dry wt of tissue; n = 6) than that in 0M females (0.05 +/- 0.00 nmol/mg dry wt; n = 8). In contrast, EGF concentration did not differ significantly between the glands of 2M and 0M males (0.51 +/- 0.01 and 1.10 +/- 0.42 nmol/mg dry wt, respectively). EGF levels were also determined in the submandibular glands from adult animals of unknown intrauterine position which were gonadectomized and then treated with testosterone and estradiol. The concentrations of EGF in the glands of gonadectomized males and females were similar (0.13 +/- 0.01 and 0.23 +/- 0.09 nmol/mg dry wt, respectively). However, there was a significant difference in response to hormonal administration between males and females. The response of females exceeded that of males at 400 and 800 micrograms testosterone/day. These results suggest that the hormonal environment of a fetus, specifically modification of the fetal environment by the production of hormones from adjacent fetuses, is a major factor in the adult expression of testosterone-responsive proteins such as EGF.

Amniotic Fluid↗
PubMed1984

Binding sites for epidermal growth factor in human uterine tissues and leiomyomas.

There is no published data regarding whether human uterine tissues and leiomyomas contain binding sites for epidermal growth factor (EGF). The present study was undertaken with 33 myometria, 13 leiomyomas, and 4 endometria. All myometria (fundus) and leiomyomas and 3 of 4 endometria specifically bound 125I-labeled mouse EGF (myometria: mean, 1.1; range, 0.1-3.9 fmol/mg protein; leiomyomas: mean, 1.1; range, 0.2-2.6 fmol/mg protein; endometria: mean, 1.0; range, 0.0-3.1 fmol/mg protein). [125I]EGF binding to myometrium was ligand specific in that only unlabeled EGF, but not unlabeled insulin, hCG, human PRL, prostaglandin E1, or prostaglandin F2 alpha, competed with [125I]EGF for binding. The apparent dissociation constants and specific binding capacities for myometrium and leiomyoma were: 0.7 nM, 1.9 fmol/mg protein; and 0.1 and 3.7 nM, 0.1 and 4.4 fmol/mg protein, respectively. While smooth muscle content decreased from the fundus to the cervical end of uteri, [125I]EGF binding did not correspondingly decrease (r = 0.5; n = 4). The binding of [125I]EGF to myometrium did not vary with the phase of the menstrual cycle or the patients' diagnosis before hysterectomy (P greater than 0.1). In summary, these results demonstrate that human uteri and leiomyomas contain specific, high affinity binding sites for EGF, but the binding neither exhibited topographical changes nor varied with the phase of the menstrual cycle or benign pathological state of the tissue.

Adult↗
PubMed1984

Binding sites for epidermal growth factor in human fetal membranes.

The present study demonstrates that human fetal membranes bind 125I-epidermal growth factor (125I-EGF), with chorion binding more than amnion. The chorion binding was also higher than that by decidua, but lower than in placenta. The lower binding by chorion compared to placenta was entirely attributable to a lower number of available EGF receptors and not to lower affinity. Chorion and placenta , but not amnion or decidua, from Cesarean section bound significantly more 125I-EGF when compared to tissues obtained after vaginal delivery. The binding of 125I-EGF to chorion exhibited dependency on time, temperature of incubation, pH of the incubation media, and amount of chorion protein. The 125I-EGF was not degraded during the binding reaction with chorion. The binding was specific in that unlabeled EGF inhibited 125I-EGF binding in a dose-dependent manner, whereas very high concentrations of other unlabeled hormones and growth factors had minimal effects on 125I-EGF binding. When some of these other hormones were tested for binding as tracers (125I-hCG, [3H]prostaglandin E1 and F2 alpha), neither chorion, amnion, decidua, nor placenta specifically bound these ligands. The 125I-EGF specific binding to chorion was saturable and a Scatchard plot of this data was curvilinear, which appears to be due to negative cooperativity. The apparent dissociation constant calculated from the initial slope of the plot was 0.20 nM, which is in excellent agreement with the concentration of unlabeled EGF required for half maximal inhibition of 125I-EGF binding, 0.26 nM. Autoradiography at the light microscope level revealed the presence of silver grains in amnion and chorion only when excess unlabeled EGF addition was withheld. The quantification of grains revealed the presence of a significantly (P less than 0.01) greater number of grains in chorion than in amnion, supporting the conclusion of the binding data. The physiological significance of EGF binding to fetal membranes and decidua is not known, but the considerable amount of EGF binding reported here suggests that they are target tissues for EGF.

Amnion↗
PubMed1984

Characterization of epidermal growth factor receptor and action on human breast cancer cells in culture.

Epidermal growth factor (EGF) may play a role in regulating growth of breast cancer cells in vivo. We have examined the action of EGF on breast cancer cells in vitro and characterized the EGF receptor as a model system for its action in vivo. All of the fourteen breast cancer cell lines which grow attached to culture dishes specifically bound EGF, including one purportedly normal breast line (HBL-100). The one cell line examined which grows as a suspension, DU-4475, did not express measurable levels of EGF binding. The number of EGF binding sites per cell for the different cell lines varied from 200 EGF binding sites/cell (for MDA-MB-436) to 700,000 EGF binding sites/cell (for MDA-MB-231), with most cell lines having approximately 10,000 EGF binding sites/cell. Scatchard analysis of EGF binding to four of the breast cell lines indicated a single class of high-affinity binding sites for MDA-MB-231 cells (Kd = 200 pM; n = 220 fmol of EGF bound/mg of cell protein); and for T-47D cells (Kd = 4 nM, n = 85 fmol of EGF bound/mg of cell protein) and curvilinear plots for MCF-7 cells and HBL-100 cells. The EGF binding to MDA-MB-231 cells was specific for EGF and was maximum after 2 hr at 37 degrees, followed by a progressive loss of cell-associated radio-activity, which was prevented by the action of the lysosomal inhibitory agent chloroquine. Specific covalent binding of 125I-EGF to MDA-MB-231 cells indicated that the EGF receptor had molecular weights of 165,000 and 140,000. MCF-7 cells and T-47D cells grown in serum-free medium supplemented with 10 nM EGF for 3 days had significantly increased protein, DNA, and cell number, whereas MDA-MB-231 and ZR-75-1 cells did not respond significantly to EGF. These results indicate that EGF receptors are consistently expressed by breast cells grown attached to a surface but that some cell lines expressing EGF receptors do not respond mitogenically to EGF. The biochemical characteristics of EGF receptors in MDA-MD-231 breast cells are similar to those observed for EGF receptors in other human tissues.

Breast Neoplasms↗
PubMed1984

Epidermal growth factor binding by breast tumor biopsies and relationship to estrogen receptor and progestin receptor levels.

Epidermal growth factor (EGF) may be important in regulating the growth of some breast cancer cells in vivo because of its mitogenic action on some breast cancer cell lines in vitro. Epidermal growth factor receptors (EGF-R) were measured in a series of breast tumors to determine what percentage of breast tumors express EGF-R and whether EGF-R was independent of expression of estrogen receptor and progestin receptor. Specific binding of 125I-EGF to membranes from pooled homogenates of breast tumors reached equilibrium after 45 min at 25 degrees and remained constant. Scatchard analysis of 125I-EGF binding indicated a single class of receptors with an apparent Kd of 2 nM and a binding capacity of 28 fmol/mg of membrane protein, and the binding of 125I-EGF was not effectively competed for by insulin, fibroblast growth factor, growth hormone, or prolactin. Specific binding of 125I-EGF of 1 fmol or greater/mg of membrane protein and 15% or greater specific binding was detected in 48% of 137 unselected primary and metastatic breast tumors. The frequency distribution of EGF binding values was unimodal, with a progressive decrease in the proportion of patients with high EGF binding values. The values of EGF binding ranged from 1 to 121 fmol/mg of protein, with an arithmetic mean of 8.4 fmol/mg of protein and a geometric mean of 3.2 fmol/mg of protein. Forty-two % of 24 metastatic breast tumors were positive for EGF binding, with an arithmetic mean of 6.3 fmol/mg of protein and a geometric mean of 4.1 fmol/mg of protein. The magnitude of EGF binding in individual tumors was independent of either estrogen receptor or progestin receptor levels, although the highest quantities of EGF binding were expressed by tumors lacking steroid receptors. Approximately 20% of the tumors in the study were EGF-R-positive and ER-negative, suggesting that the growth of these tumors may be regulated predominantly by a peptide hormone (EGF) rather than a steroid hormone (estrogen). EGF binding did not correlate significantly with age of the patients. Correlation analysis between EGF binding and the percentage of malignant and nonmalignant cell types present in sections of tumor adjacent to the area assayed for EGF binding indicated that the percentage of malignant cells is an important factor in determining the amount of EGF binding in tumor homogenates. The recent discovery of transforming growth factors which interact with the EGF-receptor system suggests additional roles for EGF receptors in breast cancer.

Binding, Competitive↗
PubMed1981

Biological activity of an angiotensin II--ferritin conjugate on rabbit aortic smooth muscle cells.

Specific binding sites for [Asp1,Ile5]angiotensin II (angiotensin) have been demonstrated in homogenates and subcellular fractions of aortic medial smooth muscle cells, but the localization of the angiotensin receptor responsible for contraction has not been determined [Devynck, M. A., & Meyer, P. (1976) Am. J. Med. 61, 758-767]. To establish the location of this receptor, we have prepared a membrane-impermeable analogue of angiotensin by acylating its N-terminal amino group with the N-hydroxysuccinimide ester of succinylated ferritin. This angiotensin-ferritin conjugate possessed the same intrinsic activity as angiotensin but was approximately 200 times less potent in inducing contraction in rabbit aortic strips. The stability of the conjugate was investigated, and approximately 5% of the contractile activity of the angiotensin-ferritin conjugate was attributable to low molecular weight components that were present before or after exposure to aortic strips. The time required for aortic strips to reach a plateau of contraction in response to angiotensin-ferritin was significantly longer than that required by free angiotensin to produce the same level of contraction. With enzymatically dispersed aortic smooth muscle cells, however, the time taken to produce contractions by both angiotensin and angiotensin-ferritin was indistinguishable. [Sar1,-Ala8]angiotensin II, a competitive inhibitor of angiotensin, completely suppressed contractions induced by angiotensin or angiotensin-ferritin in aortic strips or dispersed aortic smooth muscle cells. These results suggest that angiotensin need not directly penetrate the plasma membrane to cause contraction and imply that the angiotensin receptor responsible for initiating contraction of aortic smooth muscle cells is located on the plasma membrane.

Angiotensin II↗
PubMed1978

Preparation of functional smooth muscle cells from the rabbit aorta.

A procedure for dissociating the rabbit aorta into single, functional smooth muscle cells is described. After removal of adventitia and intima, slices of media were incubated with purified collagenase, elastase, and soybean trypsin inhibitor in a Krebs-Ringer buffer modified with Hepes, amino acids, and a [Ca2+] of 0.2 mM. After enzymatic digestion and mechanical shear, the yield of dispersed cells was approximately 25% based on DNA recovered. Greater than 95% of the cells excluded trypan blue and approximately 80-90% adhered to tissue culture dishes. By phase contrast microscopy, most of the cells were elongate and approximately 10 micron X 30 micron in size. The remainder were either spherical or highly crenated and contracted. Electron microscopy of the cells showed that immediately after dissociation greater than 95% could be identified as smooth muscle, though most had undergone some degree of structural change compared to cells in situ. Depending on the preparation, from 5 to 50% of these cells contracted in response to agonists. Cells shortened by 10-15% and developed numerous evaginations when stimulated by angiotensin II norepinephrine, or carbamylcholine. Cells relaxed after washout of agonists and could subsequently be restimulated. Specific inhibitors of each of the agonists blocked the contractile response. Dispersed cells cultured for 1-5 days contracted in even higher numbers than the freshly prepared cells, suggesting restoration of hormone binding and/or contractile function in culture. This preparation provides a system in which the physiology of individual vascular smooth muscle cells may be studied.

Animals↗
PubMed1977

Competition of delta 9-tetrahydrocannabinol with estrogen in rat uterine estrogen receptor binding.

Direct competition experiments with delta 9 -tetrahydrocannibinol (delta 9-THC) and estradiol in binding assays with rat uterine cytosol estrogen receptors showed that delta 9-THC was a weak, but nevertheless significant, competitor for binding to cytoplasmic estrogen receptors. These data support, at the molecular level, the observations that delta 9-THC has a weak estrogenic activity (at least the ability to bind to estrogen receptors). Moreover, estrogen-like binding suggests that delta 9-THC, acting at the level of estrogen receptor, causes a primary estrogenic effect rather than an indirect or secondary phenomenon.

Animals↗
PubMed1977

alpha-Lactalbumin levels in human mammary tumors, sera, and mammary cell culture lines.

Antiserum to purified human alpha-lactalbumin was produced in rabbits and used to develop a radioimmunoassay capable of detecting 0.1 ng of alpha-lactalbumin per ml of sample. Human breast diseases were analyzed for alpha-lactalbumin levels. A high percentage of breast carcinomas contained varying levels of alpha-lactalbumin. Lymph node metastases from primary carcinomas that synthesized alpha-lactalbumin also contained it. Analysis of serum from breast cancer patients indicated that approximately 25 percent had measurable levels of alpha-lactalbumin before surgery, but no alpha-lactalbumin was found in postsurgery sera. alpha-Lactalbumin was not detected in the urine of early lactational women, although it was present in the sera. Human cell culture lines derived from pleural effusions of mammary carcinomas contained little, if any, alpha-lactalbumin. Other human cell lines derived from mammary carcinomas and grown as solid tumors in athymic mice did not contain measurable levels of alpha-lactalbumin.

Breast Neoplasms↗
PubMedDate not supplied

Effects of transforming growth factor-beta1 on extracellular matrix gene expression by human fibroblasts from a laryngeal stenotic lesion.

Transforming growth factor-beta1 appears to play important roles in normal wound healing by increasing synthesis of extracellular matrix components. However, the role of transforming growth factor-beta1 in the production of excessive scar tissue by fibroblasts from stenotic lesions of the larynx has not been evaluated. We examined the effect of transforming growth factor-beta1 on the steady-state messenger RNA levels of elastin, alpha2(l) procollagen, and lysyl oxidase (the enzyme that cross-links both of these structural proteins) in cell cultures of diploid human fibroblasts established from fetal skin, newborn foreskin, and an adult laryngeal stenotic lesion. Time-course and dose-response experiments demonstrated that treatment with 500 pmol/L transforming growth factor-beta1 for 20 hours induced maximal levels of mRNA for elastin (7- to 59-fold) and alpha2(l) procollagen (1.7- to 2.4-fold) in all three cultures of fibroblasts. Transforming growth factor-beta1 also increased levels of lysyl oxidase mRNA in fibroblasts cultured from newborn foreskin (2.4-fold) and a stenotic lesion (10-fold) but had minimal effects on the fibroblasts cultured from fetal skin (1.1-fold), which constitutively expressed high levels of lysyl oxidase mRNA. Furthermore, the fibroblast culture established from a laryngeal stenotic lesion responded with the highest fold-induction for all three mRNAs. Inhibition of mRNA synthesis by actinomycin D showed that transcription was required for transforming growth factor-beta1 induction of elastin, alpha2(l) procollagen, and lysyl oxidase mRNA in all three cultures of fibroblasts. Inhibition of protein synthesis by cycloheximide showed that translation was required for maximal induction by transforming growth factor-beta1 of elastin mRNA but had no observable effect on alpha2(l) procollagen mRNA in all three cultures of fibroblasts. In addition, translation was required for maximal induction of the lysyl oxidase mRNA by transforming growth factor-beta1 in the fibroblasts cultured from a stenotic lesion but not for fibroblast cultures established from fetal and adult skin. These results show that transforming growth factor-beta1 coordinately increases mRNA levels for the structural extracellular matrix proteins collagen and elastin, as well as for the cross-linking enzyme, lysyl oxidase. These data also support the hypothesis that transforming growth factor-beta1 may contribute to the formation of laryngeal stenotic lesions.

Journal Article↗
PubMedDate not supplied

Biochemical analysis of acute and chronic wound environments.

The process of wound healing involves a complex interaction between numerous cell types, extracellular matrix molecules, and soluble mediators including growth factors and cytokines. This complex milieu is under active investigation for the purposes of beginning to understand how this environment regulates tissue repair. Quantitation of growth factors, cytokines, and matrix metalloproteinases within surgical wound fluids may help to elucidate this regulatory network, not only in noncomplicated wound healing but also in pathologic lesions such as chronic ulcers.

Journal Article↗
PubMedDate not supplied

Alpha-smooth muscle actin expression in rat and mouse mesenteric wounds after transforming growth factor-beta1 treatment.

Rat mesenteric perforations heal by contraction within 5 to 7 days, whereas mouse mesenteric perforations seldom close within 3 weeks unless stimulated by transforming growth factor-beta1. In this article, we quantified the expression of alpha-smooth muscle actin by quantitative-reverse transcription-polymerase chain reaction and the orientation of actin filaments at the wound margin by Fourier transformation image analysis after treatment with transforming growth factor-beta1. The expression of transforming growth factor-beta1 and its type II receptor was also assessed. Actin filaments were shown to increase with time at the wound margin in both species and the expression of alpha-smooth muscle actin mRNA increased simultaneously. Transforming growth factor-beta1 enhanced the alpha-smooth muscle actin expression four to five times in rats and three to four times in mice on day 5, but the number of copies expressed per cell was 15-fold higher in rats than in mice. Transforming growth factor-beta1 was down-regulated after wounding in free peritoneal cells of rats, but maintained until day 5 in transforming growth factor-beta1-treated mice. The main finding of this study was that untreated, normal rats expressed substantially more alpha-smooth muscle actin than mice. After treatment with transforming growth factor-beta1, this expression increased similarly in both species. It can be hypothesized that normal closure of mesenteric perforations requires a minimum level of actin expression. This level is not reached in normal mice, but is exceeded after stimulation. Perforations in the rat always close, because the alpha-smooth muscle actin expression is always above this level.

Journal Article↗
PubMedDate not supplied

Molecular and mechanistic validation of delayed healing rat wounds as a model for human chronic wounds.

The purpose of this study was to provide molecular and mechanistic evaluation of an ischemic wound model in rats to determine if it is a valid model for human chronic wounds. Compared to acute wounds, human chronic wounds contain markedly elevated levels of proinflammatory cytokines and matrix metalloproteinases, while matrix metalloproteinase inhibitors and growth factor activity are diminished. Accordingly, tissue from ischemic and normal rat wounds were analyzed for cytokine, proteases and growth factor levels. Dorsal full thickness punch wounds were created in rats using a reproducible template. The ischemic wound group (n = 10) had six uniformly placed wounds within a bipedicled dorsal flap. The control group (n = 10) had the same wounds created without elevation of a flap. On postwound days 3, 6 and 13 wounds were excised and analyzed. Protein levels for tumor necrosis factor-alpha were determined with a rat-specific enzyme-linked immunosorbent assay, while mRNA was determined by RNase protection assay. Matrix metalloproteinases and serine protease detection was done using gelatin and casein zymography, respectively. Significant delay in healing was achieved in the ischemic group: 50% healing for control wounds was at 7 days and 11 days for ischemic wounds (p < 0.001). No significant differences between wound groups were found for interleukin-1beta, and mRNA for tumor necrosis factor-alpha and interleukin-1beta. However, at day 13 ischemic wounds contained significantly more tumor necrosis factor-alpha than controls and normal skin (586 +/- 106 pg/biopsy vs. 79 +/- 7 pg/biopsy vs. 52 +/- 2 pg/biopsy; p < 0. 001). Zymography showed substantially greater quantities of matrix metalloproteinase-2, matrix metalloproteinase-9, and serine proteases in ischemic wounds. This model of delayed healing in rats shares many of the key biochemical, molecular and mechanistic characteristics found in human chronic wounds, namely elevated tumor necrosis factor-alpha and protease levels. As such, this model will likely prove to be useful in chronic wound research, particularly in developing novel therapeutics.

Animals↗
PubMedDate not supplied

Profiles of matrix metalloproteinase activity in equine tear fluid during corneal healing in 10 horses with ulcerative keratitis.

OBJECTIVE: Levels of tear film matrix metalloproteinases (MMPs) activity are significantly elevated in horses with ulcerative keratitis and contribute to the excessive breakdown of stromal collagen. Changes in the amount of proteolytic activity in horse tear film during corneal healing and stromal remodeling have not yet been reported, but we hypothesize they should decrease. In the present study we analyzed serial tear fluid from horses with ulcerative keratitis to identify any changes in MMP activity during corneal healing and stromal remodeling. PROCEDURES: Samples of tear fluid were obtained from both eyes of 10 horses with ulcerative keratitis on the day of admission (day 1) at the hospital and then at various time points until complete healing of the cornea. Tear film MMP2 and MMP9 activity was determined by quantitative gelatin zymography. In all cases medical treatment included topical applications of equine serum, antibiotics, atropine and systemic administration of anti-inflammatory drugs. Surgical procedures were performed in several cases on day 2 in addition to the medical treatment. RESULTS: The mean total MMP activity (+/- SD) measured in relative standard units (RSU) in the tear fluid of the ulcerated eye (2.44 +/- 1.44) of the 10 horses was significantly higher than the mean in the contralateral eye (0.81 +/- 0.68) (P = 0.006), on the day of admission at the VMTH. The mean MMP activity in these ulcerated eyes significantly decreased (-82.4%) between the first day of admission and the day when the ulcer had completely healed (P = 0.0002). The activity level in the healed eye (0.43 +/- 0.17) was not significantly different to the one in the contralateral eye (0.36 +/- 0.18) on the day of complete corneal healing (P = 0.374). The level of MMP activity in the contralateral eye also decreased from 0.81 +/- 0.68-0.36 +/- 0.18 but this decrease (56%) was not significant (P = 0.069). CONCLUSIONS: Ulcerative keratitis in horses is associated with initially high levels of tear film proteolytic activity that decrease as the ulcers heal. The success of medical and surgical treatment of the corneal ulcers is reflected by the enzyme activity in tears. In horses successful treatment does lead to a rapid reduction in tear film proteolytic activity that corresponded with the improvement in the clinical signs of corneal ulceration. Measurement of MMP activity in the tear film might represent a way to monitor the progression of corneal healing in horses with ulcerative keratitis.

Animals↗