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Biomedical subjects

M K Collins

Publications and source records attributed to M K Collins.

At least 55 records · Page 3Linked to original sources
PubMed1994

Disregulation of Myc expression in murine bone marrow cells results in an inability to proliferate in sub-optimal growth factor and an increased sensitivity to DNA damage.

Derivatives of the murine bone marrow-derived, IL-3-dependent cell line, BAF3, were isolated following recombinant retroviral infection which showed disregulated expression of human c-Myc oncoprotein. Such cells entered apoptosis and lost viability more rapidly than parental BAF3 cells when IL-3 was removed. c-Myc disregulation also resulted in an inability of BAF3 cells to survive in either sub-optimal concentrations of IL-3, or saturating concentrations of IL-4 and insulin-like growth factor-1. Furthermore, BAF3 cells with disregulated c-Myc expression were more sensitive to the induction of apoptosis by DNA-damaging agents. These effects of c-Myc disregulation could be inhibited by co-expression of the Bcl-2 oncoprotein. The implications of these data for the transformation of haematopoietic cells by disregulation of Myc expression are discussed.

Animals↗
PubMed1994

Type C retrovirus inactivation by human complement is determined by both the viral genome and the producer cell.

The inactivation of type C retroviruses by human serum may be a considerable impediment to the use of retroviral vectors in vivo for gene therapy. Here we show that virus inactivation is dependent both on the virus and on the cell line used to produce the virus. All viruses produced from murine NIH 3T3 or dog Cf2ThS+L- cells are sensitive to human serum. In contrast, those produced from mink Mv-1-Lu and human HOS or TE671 cells are at least partially resistant, with the exception of murine leukemia viruses. In particular, the feline endogenous virus RD114 is completely resistant to a panel of eight human sera when produced from Mv-1-Lu or HOS cells. This differential resistance is controlled by the viral envelope proteins. Virus inactivation can be correlated with the ability of the producer cells to be lysed by human serum. Inactivation of sensitive viruses requires the classical pathway of complement but does not require virion lysis.

3T3 Cells↗
PubMed1994

Vulvar application of lidocaine for pain relief in spontaneous vaginal delivery.

OBJECTIVE: To evaluate the effect of topical perineal lidocaine on immediate postpartum perineal pain. METHODS: Two hundred three volunteers randomly received either 2% lidocaine jelly or chlorhexidine gluconate topically to the perineum during the second stage of labor in a double-blind study. A four-point analogue pain scale was used to rate perineal pain 30 minutes after delivery. Parametric data were evaluated with the unpaired Student t test. Nonparametric data were analyzed using Mann-Whitney, Pearson chi 2, and Mantel-haenzel tests, and forward stepwise logistic regression. RESULTS: Women receiving topical lidocaine reported less overall perceived pain (48%, P < .05) and less moderate to severe pain (52%, odds ratio 1.83, P < .04) at delivery than those receiving placebo (33 and 67%, respectively). The incidence of perineal lacerations was similar in the two groups. A logistic regression found topical lidocaine and multiparity as significant correlates of diminished peripartum pain (P < .007). CONCLUSIONS: Topical application of 2% lidocaine gel was associated with decreased pain perception in the immediate postpartum interval. If confirmed by other investigators, this technique may offer improved analgesia while minimizing the injection route with local analgesia.

Administration, Topical↗
PubMed1994

Growth factor starvation of bcl-2 overexpressing murine bone marrow cells induced refractoriness to IL-3 stimulation of proliferation.

Murine bone-marrow derived BAF3 cells, over-expressing the human Bcl-2 gene product, showed considerably delayed onset of apoptosis when deprived of IL-3. Such Bcl-2-BAF3 cells arrested rapidly in the G1 phase of the cell cycle upon IL-3 removal, then became refractory to IL-3 re-stimulation. The delay in IL-3 induced proliferation of Bcl-2 over-expressing cells was due to down-regulation of a specific signalling pathway. In the refractory cells, IL-3 was able to stimulate protein tyrosine phosphorylation and c-myc mRNA accumulation, but not rapid Erk2 activation or cdc2 mRNA accumulation.

Animals↗
PubMed1993

Immunization with interleukin-2-secreting allogeneic mouse fibroblasts expressing melanoma-associated antigens prolongs the survival of mice with melanoma.

The survival of C57BL/6 mice (H-2b) bearing B16 melanoma (H-2b) was prolonged if the animals were treated solely by immunization with an interleukin-2 (IL-2)-secreting allogeneic cell construct that expressed melanoma-associated antigens (MAA) along with major histocompatibility complex (MHC) class-I determinants (H-2k; RLBA-IL-2 cells). This was the case if the mice were immunized simultaneously with, or 6 days following, the injection of viable B16 cells. Under similar conditions, the survival of tumor-bearing mice immunized with an allogeneic cell construct (H-2k) that expressed MAA but did not secrete IL-2 (RLBA-ZipNeo cells), or with an allogeneic construct (H-2k) that secreted IL-2 but did not form MAA (LM-IL-2 cells), was also prolonged. However, in these instances, the period of survival was significantly shorter than that of mice immunized with the cell construct that combined IL-2 secretion with the expression of MAA. Tumor-bearing mice immunized with non-transfected LM(TK-) cells (H-2k), or irradiated B16 cells (H-2b) failed to survive longer than untreated mice. Although the survival of the treated animals was prolonged, in most instances tumor growth recurred. The recurrent tumors in mice treated with the allogeneic cell constructs formed melanin and were histologically indistinguishable from tumors in untreated mice. Cells from the recurrent tumors were resistant to further immunotherapy and to cytotoxic effector cells obtained from the spleens of mice immunized with the same cellular immunogen used initially. The injection of IL-2-secreting syngeneic B16 cells into C57BL/6 mice invariably resulted in the appearance of non-IL-2-secreting melanomas. Under similar circumstances, tumors failed to develop in C57BL/6 mice injected with IL-2-secreting, or non-secreting, allogeneic cell constructs. Thus, the expression of allogeneic antigens protected the mice from growth of the cellular immunogens.

Animals↗
PubMed1993

Interleukin 2 activates extracellular signal-regulated protein kinase 2.

Interleukin 2 (IL-2) stimulated activation of the 42-kD extracellular signal-regulated kinase 2 (Erk2) in murine IL-3-dependent cells, expressing either high or intermediate affinity IL-2 receptors. Activation was both rapid, occurring within 5 min of IL-2 addition, and prolonged, remaining elevated for 30 min. Activation of Erk2 appeared to be necessary for IL-2 stimulation of proliferation, as deletion of a region of the cytoplasmic domain of the IL-2 receptor beta chain, essential for IL-2 stimulation of proliferation, abolished Erk2 activation by IL-2. Furthermore, cells that had been deprived of cytokine for 24 h were then refractory to IL-2 stimulation of both Erk2 activity and proliferation. However, elevation of Erk2 activity was not sufficient to stimulate proliferation, as protein kinase C activation stimulated Erk2 activity but not DNA synthesis. Also, cells exposed to IL-2 in the presence of rapamycin showed full Erk2 activation but not DNA synthesis. These data suggest that IL-2 must stimulate both Erk2 activity and a further pathway(s) to trigger cell proliferation.

Animals↗
PubMed1993

X-linked chronic granulomatous disease: correction of NADPH oxidase defect by retrovirus-mediated expression of gp91-phox.

Chronic granulomatous disease (CGD) is an inherited immunodeficiency resulting from the inability of an individual's phagocytes to produce superoxide anions because of defective NADPH oxidase. The disease may be treated by bone marrow transplantation and as such is a candidate for somatic gene therapy. Two thirds of patients have defects in an X-linked gene (X-CGD) encoding gp91-phox, the large subunit of the membrane cytochrome b-245 component of NADPH oxidase. Epstein-Barr virus-transformed B-cell lines from patients with CGD provide a model system for the disease. We have used retrovirus-mediated expression of gp91-phox to reconstitute functionally NADPH oxidase activity in B-cell lines from three unrelated patients with X-CGD. The protein is glycosylated and membrane associated, and the reconstituted oxidase is appropriately activated via protein kinase C. The kinetics of superoxide production by such reconstituted cells is similar to that of normal B-cell lines. These data show the potential of gene therapy for this disease.

B-Lymphocytes↗
PubMed1993

Bcl-2 oncogene protects a bone marrow-derived pre-B-cell line from 5'-fluor,2'-deoxyuridine-induced apoptosis.

The bcl-2 protooncogene has been shown to protect haemopoietic precursors from programmed cell death after the removal of interleukin-3 (IL3). In the present report we show evidence that overexpression of bcl-2 in the pre-B-cell line BAF3 protects cells from apoptosis induced by treatment with the thymydilate synthase inhibitor 5'-fluor,2'-deoxyuridine (FDUR) in the presence of IL-3. Dose-response experiments analyzing the dependence of cell death on drug concentration indicated a marked resistance of BAF3bcl-2 to FDUR treatment. Cleavage of DNA into oligonucleosome-length fragments, a characteristic of apoptosis, was observed in BAF3 cells and inhibited in the cells overexpressing bcl-2. We have determined variations in the dATP and dTTP pools after FDUR treatment. Interestingly, no differences were found between both cells in the kinetics of changes in dNTP pools. Therefore, the protective effect of the Bcl-2 protein on apoptosis induced by dNTP unbalance must be ascribed to a step downstream of perturbations in the synthesis of DNA precursors and before activation of endonucleolytic cleavage of chromatin.

Animals↗
PubMed1993

Mutation of Asp20 of human interleukin-2 reveals a dual role of the p55 alpha chain of the interleukin-2 receptor.

Mutation of Asp20 in human interleukin-2 (IL-2) to Lys is known to result in an IL-2 molecule with unchanged binding to the p55 subunit of the IL-2 receptor, but with greatly decreased affinity for the p75 subunit (Collins, L., Tsien, W.-H., Seals, C. et al. Proc. Natl. Acad. Sci USA 1988. 85: 7709). Here we demonstrate that Lys20 IL-2 competed with a reduced (10-fold) affinity for high-affinity IL-2 receptors on two murine cell lines HT2 and CTLL. In parallel with this difference in receptor interaction, Lys20 IL-2 stimulated half-maximal HT2 cell proliferation at a 10-fold higher concentration than wild-type IL-2. However, half-maximal stimulation of CTLL cells required a 100-fold higher concentration of Lys20 IL-2. A similar 100-fold reduction in bioactivity of Lys20 IL-2 was observed for primary, activated, human or murine lymphocytes. Anti-p55 antibodies increased the concentration of Lys20 IL-2 required to stimulate HT2 cells to that required for CTLL cells. These data suggest that CTLL cells, while able to bind Lys20 IL-2 with high affinity, are lacking a p55-dependent function necessary for optimal stimulation. Therefore, p55 has a dual role, being important both for high-affinity IL-2 binding and for optimal cell triggering.

Animals↗
PubMed1993

The control of apoptosis in mammalian cells.

This article summarizes what is now known, more than 100 years after this prescient observation, of the mechanisms by which death of mammalian cells is regulated. Particular attention is paid to the elimination of self-reactive thymocytes, the induction of cell death during cancer therapy and the mechanism(s) by which cells die following growth factor removal.

Animals↗
PubMed1993

Comparison of the potential therapeutic effects of interleukin 2 or interleukin 4 secretion by a single tumour.

Engineering of a variety of rodent tumour cells to secrete either interleukin 2 (IL-2), or interleukin 4 (IL-4), has been demonstrated to reduce their tumorigenicity. However the mechanisms of action of secreted IL-2 and IL-4 have not been compared in a single rodent tumour. Here we demonstrate that the weakly immunogenic murine fibrosarcoma FS29 had reduced growth rate and in some cases was rejected by syngeneic animals, when modified to secrete either IL-2 or IL-4, but not IL-5. Immunohistochemical analysis of tumour nodules undergoing regression showed stimulation of a largely lymphocytic infiltrate by IL-2 and a macrophage and granulocyte infiltrate, with a small number of lymphocytes by IL-4. Indeed, secretion of low levels of IL-2 and IL-4 in combination resulted in optimal rejection, suggesting that the two cytokines might mobilise different and complementary effector cell mechanisms. Both IL-2 and IL-4-secreting cells failed to induce the rejection of admixed, unmodified FS29 cells. The loss of cytokine secreting cells from such admixtures occurred more rapidly for IL-2-secreting cells. Injection of IL-4-secreting, but not IL-2-secreting FS29 cells could protect mice from a delayed challenge with unmodified FS29 cells. These data suggest that IL-4 secretion stimulates the better long-term host anti-tumour response.

Animals↗
PubMed1993

The role of IL-2 interaction with p75 and p55 receptor molecules in the stimulation of cell proliferation.

This report addresses the role of individual IL-2 binding proteins of the IL-2 receptor in the stimulation of cell proliferation by IL-2. Murine IL-3-dependent cell lines were established which expressed the human p75 IL-2-binding protein, in the absence or presence of the human p55 IL-2-binding protein. Whereas p75 expression was sufficient to confer response to an intermediate (half-maximal stimulation at 100 pM) concentration of IL-2, additional expression of p55 increased the sensitivity of the cells to half-maximal stimulation at 10 pM IL-2. A mutant IL-2 molecule, Lys-20 IL-2 which is known to be defective of p75 interaction, was unable to stimulate cells expressing only p75: p55 co-expression could restore its activity. Under conditions of low p75 expression, Lys-20 IL-2 could act as an antagonist of wild-type IL-2 action. These data support a role for p55 in the enhancement of responsiveness to IL-2.

Animals↗
PubMed1993

Independent cell types are involved in the induction of antimelanoma responses in C57BL/6 mice immunized with interleukin-2-secreting allogeneic mouse fibroblasts expressing melanoma-associated antigens.

Cellular antimelanoma immune responses developed during the coincubation of spleen cells from naive C57BL/6 mice (H-2b) with LM mouse fibroblasts (H-2k) genetically modified to express melanoma-associated antigens (MAA) and to secrete interleukin (IL)-2 (RLBA-IL-2 cells). Antimelanoma responses also developed following coincubation of spleen cells from naive mice with allogeneic cells (H-2k) that expressed MAA, but did not secrete IL-2 (RLBA-ZipNeo cells), or allogeneic cells (H-2k) that secreted IL-2, but did not form MAA (LM-IL-2 cells). However, in these instances, the magnitude of the responses was significantly less (p < 0.01) than followed coincubation of spleen cells with the allogeneic cell construct (RLBA-IL-2) that combined IL-2 secretion with the expression of MAA. LM(TK-) cells (H-2k) or B16 cells (H-2b) failed to generate antimelanoma immune responses in populations of spleen cells from C57BL/6 mice. The cell types involved in the induction of the antimelanoma response by the genetically modified cells were determined by prior depletion of T-cell subsets with monoclonal antibodies before the addition of the cellular immunogens. Antimelanoma responses failed to develop in spleen cell populations depleted of T-helper cells if the cellular immunogen was non-IL-2 secreting (RLBA-ZipNeo cells). Prior depletion of T-helper cells did not affect the induction of an antimelanoma response in cell populations coincubated with constructs (RLBA-IL-2 or LM-IL-2) modified to secrete IL-2.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Cytokine gene transfer as a therapeutic strategy.

Cytokine gene therapy for cancer could involve either the direct delivery of cytokine genes to established tumours to stimulate their rejection or the injection of cytokine-secreting tumour cells to stimulate an immune response that could reduce metastatic disease. To assess the feasibility of the first approach, we have compared the ability of different cytokine-secreting tumour cells to induce the rejection of admixed, unmodified cells. While interleukin (IL)-2- or interleukin-4-secreting tumour cells were ineffective, interferon-gamma (IFN-gamma)-secreting cells could induce rejection of 10% admixed, unmodified cells. Because direct gene delivery to tumours is unlikely to be 100% efficient, these data suggest that IFN-gamma may be the most suitable of these cytokines for this approach. However, we have demonstrated that injection of IL-2-secreting tumour cells, following primary tumour excision, can prevent the development of metastases and prolong survival of rats. This suggests that IL-2-secreting tumour cells can be effective in the treatment of metastatic disease.

Animals↗
PubMed1993

Cell cycle arrest of proliferating neuronal cells by serum deprivation can result in either apoptosis or differentiation.

Apoptotic cell death plays a critical role in the development of the nervous system. The death of mature nondividing neurons that fail to receive appropriate input from the target field has been extensively studied. However, the mechanisms mediating the extensive cell death occurring in areas of the developing brain where proliferating neuroblasts differentiate into mature nondividing neurons have not been analyzed. We show here that the cell cycle arrest of a proliferating cell of neuronal origin by removal of serum results in either apoptotic cell death or differentiation to a mature nondividing neuronal cell. The proportion of cells undergoing death or differentiation is influenced in opposite directions by treatment of the cells with cyclic AMP and retinoic acid. This suggests that following the withdrawal of signals stimulating neuroblast cell division, neuronal cells either can cease to suppress a constitutive suicide pathway and hence die by apoptosis or, alternatively, can differentiate into a mature neuronal cell. Regulation of the balance between apoptosis and neuronal differentiation could therefore play a critical role in controlling the numbers of mature neurons that form.

Animals↗
PubMed1993

Mutation of amino acids within the gibbon ape leukemia virus (GALV) receptor differentially affects feline leukemia virus subgroup B, simian sarcoma-associated virus, and GALV infections.

The three type C retroviruses, gibbon ape leukemia virus (GALV), simian sarcoma-associated virus (SSAV), and feline leukemia virus subgroup B (FeLV-B), infect human cells by interacting with the same cell surface receptor, GLVR1. Using LacZ retroviral pseudotypes and murine cells transfected with mutant GLVR1 expression vectors, we show that the same 9-amino-acid region of human GLVR1 is critical for infection by the three viruses. Rat cells were not susceptible to infection by LacZ (FeLV-B) pseudotypes because of a block at the receptor level. We found multiple amino acid differences from human GLVR1 in the 9-amino-acid critical region of rat GLVR1. Expression of a human-rat chimeric GLVR1 in murine cells demonstrated that rat GLVR1 could function as a receptor for GALV and SSAV but not for FeLV-B. Substitution of human GLVR1 amino acids in the critical region of rat GLVR1 identified three amino acids as responsible for resistance to FeLV-B infection; two of these affect SSAV infection, but none affects GALV infection.

Amino Acid Sequence↗