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D R Twardzik

Publications and source records attributed to D R Twardzik.

At least 55 records · Page 3Linked to original sources
PubMed1985

A beta-type transforming growth factor, present in conditioned cell culture medium independent of cell transformation, may derive from serum.

An alpha-type transforming growth factor (TGF alpha) is produced at high levels by rat embryo cells transformed by the Snyder-Theilen strain of feline sarcoma virus (FeSV). Addition of 2 ng mouse epidermal growth factor (mEGF) during purification identified the presence of a second, EGF-dependent growth factor of the TGF beta type (TGF beta) in this conditioned medium. This factor had an approximate Mr of 12,000 and eluted at 37% acetonitrile during high performance liquid chromatography. This extracellular type of TGF beta activity also was present in conditioned medium of rat cells after infection with a transformation defective strain of Abelson leukemia virus, and hence expression of this growth factor activity was independent of cell transformation. Moreover, the presence of an EGF-dependent, 12,000 Mr clonogenic activity in extracts of bovine serum alone suggests serum as an origin for the B-type transforming growth factor initially observed in conditioned medium of Snyder-Theilen FeSV transformed cells. This does not, however, preclude the possibility that TGF beta is also secreted by the transformed rat embryo cells themselves.

Animals↗
PubMed1985

Transforming growth factor (TGF) activity in human urine: synergism between TGF-beta and urogastrone.

Human urine contains an acid and heat stable peptide with an apparent molecular weight of 8,000-10,000 that, in the presence of urogastrone (EGF), induces the anchorage-independent growth of nontransformed cells in semisolid media. This nonmitogenic growth-modulating activity does not compete with EGF for binding to EGF membrane receptor sites and can be resolved from EGF by high-performance liquid chromatography. The urine-derived growth factor has been purified more than 10,000-fold and shares many biochemical properties with and is functionally related to the B class of TGFs isolated from transformed cells and non-neoplastic tissues. The low molecular weight anchorage-independent growth-stimulating activity universally present in human urine is a result of the synergistic interaction of this urine-derived TGF-beta and urogastrone.

Animals↗
PubMed1985

Vaccinia virus-infected cells release a novel polypeptide functionally related to transforming and epidermal growth factors.

The recent discovery, that a vaccinia virus (VV) gene encodes a polypeptide with structural homology to transforming growth factor (TGF-alpha) and epidermal growth factor (EGF), led us to look for a virus-induced protein with the predicted biological activity. The supernatants of VV-infected cell cultures were found to contain an acid stable Mr 25,000 polypeptide that competes with EGF for binding to EGF membrane receptors. This VV-induced growth factor (VGF) like EGF and TGF-alpha is mitogenic and stimulates anchorage-independent cell growth in the presence of TGF-beta. However, VGF did not cross-react in a radioimmunoassay specific for small and large forms of TGF-alpha and exhibited minimal cross-reactivity with antisera to EGF. VGF was detectable in the culture medium within 2 hr, and maximal amounts were present 12 hr after infection. The level of VGF was proportional to the multiplicity of VV used. Inhibition of viral DNA synthesis enhanced VGF production, consistent with the hypothesis that VGF is an early gene product encoded by VV. The demonstration of a novel growth factor, released from cells infected with VV, may have important implications regarding the nature of virus-host interactions.

Animals↗
PubMed1985

Detection of larger polypeptides structurally and functionally related to type I transforming growth factor.

Peptides corresponding to various regions of type I rat transforming growth factor (rTGF) have been chemically synthesized, conjugated to carrier protein, and used to immunized rabbits. An antiserum raised against one of these peptides, corresponding to the carboxyl-terminal 17 amino acids of rTGF, has been used to develop a competitive RIA for the immunizing peptide. In this assay, the antiserum reacts only with a restricted region of the immunizing peptide corresponding to the 11 carboxyl-terminal amino acids of the rTGF molecule. Intact rTGF competes as well as the immunizing peptide on a molar basis, indicating that the cognate sequence in rTGF is recognized by this antiserum. Mouse epidermal growth factor (mEGF) was ineffective as a competitor, even at a 10,000-fold greater concentration, showing that the RIA was capable of distinguishing TGF from functionally related proteins. Several immunoreactive species were detected by gel filtration of conditioned medium from cultured retrovirus-transformed rat cells; both a low molecular weight species that corresponds to rTGF, and a higher molecular weight specie(s) copurify with biologic activity in the EGF radioreceptor assay. Immunoblotting analysis of the higher molecular weight peak fractions revealed three polypeptides (MrS, 24,000, 40,000, and 42,000) that reacted specifically with the antiserum. These findings suggest the existence of a family of larger polypeptides containing both structural and functional determinants of rTGF.

Animals↗
PubMed1985

Differential expression of transforming growth factor-alpha during prenatal development of the mouse.

Transforming growth factors (TGFs) are differentially expressed in the mouse during neonatal development. Highest levels are seen early at Day 7, and lower levels, at Day 13. Both small- and large-molecular-weight forms of TGF are found; they share many biochemical properties with rat TGF-alpha, including a similar high-pressure liquid chromatography elution profile. Although the embryo-derived activities compete with epidermal growth factor for binding to epidermal growth factor membrane receptors, they are immunologically distinct from epidermal growth factor. These embryonic polypeptides, however, do cross-react in a competitive radioimmunoassay developed using a synthetic peptide corresponding to the carboxy-17 amino acids of rat TGF-alpha as the immunogen. The highly conserved TGF-alpha family of peptides produced by some tumor cells may therefore represent derepressed forms of these embryonic growth factors. A functional role in neonatal development is proposed.

Animals↗
PubMed1985

Comparison of growth factors functionally related to epidermal growth factor in the urine of normal and human tumor-bearing athymic mice.

Urine from nude mice contains epidermal growth factor (EGF) and a minor acid-stable component with an apparent molecular weight of 20,000, which competes with EGF for binding to EGF membrane receptors and which promotes colony formation by normal rat kidney cells in soft agar. The levels of this Mr 20,000 urine-derived growth factor are increased approximately 4- to 10-fold in nude mice bearing tumors following s.c. injection of cultured human tumor cells. Following removal of the primary tumor, the concentration of this factor is reduced to basal levels, and thus, elevated levels of this growth factor appear to be dependent on tumor burden. The Mr 20,000 urinary component is separable into four EGF competing activities by high-performance liquid chromatography; the major species is immunologically related to mouse submaxillary gland EGF and therefore appears to be of host origin. However, in addition to elevated levels of host growth factor, urine from tumor-bearing mice also contains transforming growth factor activity in amounts comparable to that released by the tumor cells in culture. The tumor-derived urinary transforming growth factor activity is immunologically unrelated to EGF but is immunoreactive with an antiserum to transforming growth factor-alpha. We propose that the nude mouse may be a useful model to examine the role of both host- and tumor-derived growth factors in tumorigenesis and the usefulness of these factors as biological markers of response to therapy and tumor progression.

Animals↗
PubMed1984

Neuroblastoma cells produce transforming growth factors during exponential growth in a defined hormone-free medium.

Mouse neuroblastoma Neuro-2A cells have been cultured in a chemically defined serum-free medium consisting of a 1:1 mixture of Dulbecco's modified Eagle's medium and Ham's F-12 medium, supplemented with 30 nM selenite and 10 micrograms of transferrin per ml. In this medium, which does not contain any externally added polypeptide growth factor, cells proliferate rapidly with a doubling time of approximately equal to 10 hr. During exponential growth in this serum-free medium, Neuro-2A cells secrete a 15- to 20-kDa transforming growth factor with strong mitogenic action and the ability to induce anchorage-independent growth on nontransformed cells. This neuroblastoma-derived transforming growth factor (ND-TGF) is acid and heat stable but is sensitive to treatment with trypsin or dithiothreitol. However, it does not compete with epidermal growth factor (EGF) for receptor binding and does not require EGF receptors for its mitogenic activity. Experiments on the effects of EGF on ND-TGF-induced soft agar growth of normal rat kidney cells indicate that Neuro-2A cells secrete an EGF-potentiated TGF in addition to ND-TGF. It is suggested that Neuro-2A cells can proliferate in the absence of externally added growth factors as a result of autocrine production of polypeptide growth factors.

Animals↗
PubMed1983

Similar transforming growth factors (TGFs) produced by cells transformed by different isolates of feline sarcoma virus.

Fisher rat embryo cells transformed by each of three independent isolates of feline sarcoma virus (FeSV) are shown to release transforming growth factors (TGFs) into cell culture medium. These acid- and heat-stable peptides compete for binding to, and stimulate phosphorylation of, EGF membrane receptors and promote anchorage-independent cell growth. Cells transformed by the Gardner and Synder-Theilen strains of FeSV produce high titers of TGF (60-200 ng eq EGF/liter) while cells transformed by McDonough FeSV produce TGF at only low levels (less than 10 ng eq EGF/liter). Growth factors produced by cells transformed by each of the three FeSV isolates functionally and biochemically resemble each other, mouse sarcoma growth factor (SGF), and TGFs produced by human tumor cells.

Animals↗
PubMed1983

Transforming growth factors produced by retrovirus-transformed rodent fibroblasts and human melanoma cells: amino acid sequence homology with epidermal growth factor.

Transforming growth factors (TGFs) were purified from serum-free medium conditioned by retrovirus-transformed Fisher rat embryo fibroblasts, mouse 3T3 cells, and two human melanoma cell lines. The purification of each TGF was monitored in a radioreceptor assay based on receptor crossreactivity with mouse submaxillary gland epidermal growth factor (mEGF) and was achieved by gel permeation chromatography of the acid-soluble TGF-containing activity, followed by reverse-phase high-pressure liquid chromatography with sequential use of acetonitrile and 1-propanol in the presence of aqueous trifluoroacetic acid. The amino-terminal sequences of rat, mouse, and human TGFs were determined. Extensive sequence homology was found among TGF polypeptides from different species and cell types. Alignment of the amino acid sequences of rat TGF, mEGF, and human urogastrone (hEGF) reveals statistically significant sequence homology. The reported results suggest that TGFs that compete for binding to the cellular EGF receptor and EGF may have evolved from a common progenitor.

Amino Acid Sequence↗
PubMed1983

High-molecular-weight transforming growth factor activity in the urine of patients with disseminated cancer.

Urine from 22 patients with a variety of disseminated cancers and from an equivalent number of nonmalignant controls of similar age and sex was tested for the presence of transforming growth factor (TGF) activity as measured by the ability to promote the growth in soft agar of nontransformed indicator cells. Cancer patients included those with carcinomas of the lung, breast, colon, and ovary, as well as melanomas and sarcomas. The nonmalignant controls included both normals and individuals with a variety of inflammatory and infectious disorders. Aliquots of unfrozen urine were acid extracted, chromatographed on a Bio-Gel P-30 column, and then tested for TGF activity using normal rat kidney fibroblasts and epidermal growth factor (EGF)-competing activity with human carcinoma A431 cells. These assays revealed that a high-molecular-weight TGF activity (Mr 30,000 to 35,000) which coelutes with EGF-competing activity was present in 18 of 22 cancer patients but present in only five of 22 nonmalignant controls (p less than 0.01). In contrast, a low-molecular-weight TGF activity (Mr 6000 to 8000) which does not coelute with EGF-competing activity was found in all urines tested. These results indicate that an EGF-related, high-molecular-weight TGF activity is found in the urine of cancer patients and may be a useful tumor marker. Unlike other tumor markers described previously, high-molecular-weight TGF activity has a biological activity which is related to the expression of the transformed phenotype.

Binding, Competitive↗
PubMed1982

Transformation induced by Abelson murine leukemia virus involves production of a polypeptide growth factor.

Rat embryo fibroblasts transformed by Abelson murine leukemia virus (MuLV) produce and release a transforming growth factor (TGF). Production of this factor is correlated with a tyrosine-specific protein kinase that is functionally active and is associated with the major Abelson MuLV gene product, P120. Transformation-defective mutants of Abelson MuLV do not transform cells, do not have their virus coded transforming gene product phosphorylated in tyrosine, and do not induce TGF production. Abelson MuLV-induced TGF morphologically transforms cells in culture, competes with 125I-labeled epidermal growth factor (EGF) for binding to cell receptors, and induces phosphorylation of tyrosine acceptor sites in the 160,000-dalton EGF membrane receptor. After purification to homogeneity, Abelson virus-induced TGF migrates as a single polypeptide with an apparent size of 7400 daltons as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Abelson murine leukemia virus↗
PubMed1982

Mouse embryonic transforming growth factors related to those isolated from tumor cells.

Growth factors with the two characteristic properties of sarcoma growth factor, the ability to stimulate anchorage-independent growth of normal mouse or rat fibroblasts and the ability to compete with 125I-labeled epidermal growth factor for receptor binding, can be isolated from 12- to 13-day-old normal mouse embryos of various strains. Two size classes of the transforming factor from embryo cells can be isolated with apparent molecular weights of 20,000 and 10,000. The lower-molecular-weight factor has been purified several hundred-fold and has the same properties as the peptides with a molecular weight of 10,000 produced by mouse sarcoma virus-transformed 3T3 cells. Whole-mouse embryos contain approximately 10% as much sarcoma growth factor per g of tissue as do the murine sarcoma virus-transformed cells; how it is distributed among the embryonic tissues remains to be determined.

Agar↗
PubMed1982

Transforming growth factors in the urine of normal, pregnant, and tumor-bearing humans.

Transforming growth factor (TGF) activities could be detected in the urine of normal, pregnant, and tumor-bearing humans. These acid- and heat-stable polypeptides competed for binding to epidermal growth factor (EGF) membrane receptors and promoted the anchorage-independent growth of nontransformed rodent cells. They differed from human EGF in their apparent molecular weights and soft-agar growth-stimulating activity. The urine from pregnant females contained TGF activities with apparent molecular weight(s) (relative) (Mr) of 10,000 ad 17,000--20,000. In the case of a lung cancer patient, an additional major activity of approximately 30,000--35,000 Mr was found. All urine specimens examined also contained a "common" 8,000-Mr soft-agar growth-stimulating activity, which competed for binding to EGF membrane receptors and which was chromatographically separable from EGF (urogastrone). Thus urine may provide a convenient and readily available source for the biochemical characterization of these TGF-like activities, some of which may be clinically useful biologic markers for certain types of cancer.

Carcinoma, Small Cell↗
PubMed1979

Depression of Rauscher leukemia virus envelope glycoprotein gp71 binding by lymphoid cells during leukemogenesis in mice.

The availability of membrane receptors for the 71,000-dalton envelope glycoprotein (gp71) of Rauscher murine leukemia virus on splenic and thymic cells from BALB/c mice during Rauscher murine leukemia virus-induced leukemogenesis was determined utilizing a radiolabeled gp71 binding assay. Shortly after infection, the relative cellular [125I]gp71 binding level decreased, first with splenic cells (at day 7 to 10 after infection) and later with thymic cells (at day 10 to 20 after infection). The dependency of the reduction of binding on the replication of the inoculated virus was demonstrated by regression analyses using cellular gp71 binding level as the dependent variable and infectious virus titer, as well as viral gp71 and p30 levels, of spleens and thymuses from infected mice as independent variables. With each independent variable, the reduction of gp71 binding for both cell types was highly dependent (P less than 0.01) on the level of virus detected in their respective organ. In the early stages of leukemogenesis, the [125I]gp71 binding level declined to approximately 20 to 30% of control values. During this period the rate of reduction of binding was very rapid and, in general was similar for both splenic and thymic cells. Further progression of the disease resulted in little or no further reduction in binding. The application of this technique to monitor host ecotropic virus synthesis and to study cell surface virus receptor control mechanisms in vivo is discussed.

Animals↗