Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effect was studied of pretreatment with concanavalin A (ConA) of primary myoblast cultures on their migration when transplanted into muscles. As donors, transgenic CD1 mice in which the beta-galactosidase gene is under the control of a CMV promoter (CMVLacZ.9) were used. The myoblasts were grown with 20 microg/mL ConA during the 2 days before injecting them in the right tibialis anterior (TA) muscles of BALB/c mice and mdx mice. As a control, myoblasts from the same primary cultures were grown without ConA and injected in the left TA muscles. The host muscles were not previously irradiated or damaged by notexin injection. The recipient mice were immunosuppressed with FK506. Four days after myoblast transplantation, the area occupied by donor cells was significantly greater (more than threefold) following culture with ConA than without ConA. This result indicates that culture of myoblasts with ConA permits them to migrate farther following their transplantation in host muscles not previously damaged by notexin injection or irradiation. This suggests that pretreatment with ConA may be helpful for myoblast transplantation in humans. The mechanism of this effect still remains to be investigated.
 ...
PMID:Prior culture with concanavalin A increases intramuscular migration of transplanted myoblast. 948 57

Dog myoblasts obtained from muscle biopsies were infected in vitro with a defective retroviral vector containing a cytoplasmic beta-galactosidase (beta-Gal) gene. These myoblasts were initially transplanted in the irradiated muscles of SCID mice and beta-Gal positive muscle fibers were observed. beta-Gal myoblasts were also transplanted back either in the donor dogs (autotransplantation model) or in unrelated recipient dogs (allotransplantation model). Following these myoblast injections, a rapid inflammatory reaction developed within the muscle as indicated by an expression of P-selectin and of pro-inflammatory cytokine mRNAs (interleukin 6 (IL-6) and transforming growth factor beta (TGF-beta), and by a neutrophil infiltration. Following either auto- or allotransplantation in inadequately or non-immunosuppressed dogs, a specific immune reaction also developed within 2 weeks as indicated by the infiltration of CD4+ and of CD8+ lymphocytes, the increased expression of IL-10 and granzyme B mRNAs and the presence of antibodies reacting with the injected cells. Some dogs were immunosuppressed with several combinations of FK506, cyclosporine (CsA) and RS-61443. In dogs immunosuppressed with CsA combined with RS-61443, only a few myoblasts and myotubes expressing beta-Gal were observed 1-2 weeks after the transplantation, but no muscle fibers expressing beta-Gal were observed after 4 weeks, and antibodies against the injected cells were formed. In dogs immunosuppressed with FK506 alone, although no antibodies against the injected cells were produced, there were no small cells and no muscle fibers expressing beta-Gal 1 month after the transplantation. However, FK506 triggered diarrhea and vomiting in dogs. When the dogs were immunosuppressed with FK506 combined with CsA and RS-61443, muscle fibers expressing beta-Gal were present 4 weeks after the transplantation and no antibodies reacting with donor myoblasts were detected. These results indicate that the combination of three immunosuppressive agents (i.e., FK506, CsA and RS-61443) is effective in controlling the specific immune reactions following myoblast transplantation in dogs and they underline that the outcome of myoblast transplantation is dependent in part on an adequate immunosuppression. These results obtained here in normal dogs may justify myoblast transplantation in dystrophic dogs despite the side effects of FK506.
 ...
PMID:Myoblast transplantation in non-dystrophic dog. 960 63

AtFKBP12 is an Arabidopsis cDNA that encodes a protein similar to the mammalian immunophilin, FKBP12. AtFKBP12 was used as 'bait' in a yeast 2-hybrid system to screen for cDNAs in Arabidopsis encoding proteins that bind to FKBP12. Two partial cDNAs were recovered encoding the C-terminus of a protein we have called Arabidopsis thaliana FKBP12 interacting protein 37 (AtFIP37). AtFIP37 is similar to a mammalian protein, FAP48, that also binds to FKBP12. The interaction between AtFKBP12 and AtFIP37 in the 2-hybrid system, as assessed by histidine auxotrophy and beta-galactosidase activity, was disrupted by FK506, but not by cyclosporin A, a drug that binds to cyclophilin A. AtFIP37 was also shown to bind in vitro to AtFKBP12 in GST-fusion protein binding assays. The binding was abolished by prior incubation of AtFKBP12 with FK506. These findings indicate that an Arabidopsis FKBP12 ortholog encodes a protein that binds FK506 and that the interaction between AtFKBP12 and AtFIP37 may involve the FK506 binding site of AtFKBP12. The interaction provides interesting new opportunities for controlling protein:protein interactions in vivo in plants.
 ...
PMID:An Arabidopsis immunophilin, AtFKBP12, binds to AtFIP37 (FKBP interacting protein) in an interaction that is disrupted by FK506. 980 17

Mutations in the gene encoding for the myelinating Schwann cell protein P0 have been linked to inherited peripheral neuropathies, including the Charcot-Marie-Tooth type 1B disease (CMT1B) and Dejerine-Sottas syndrome (DSS). Recently generated mice deficient in the P0 gene (P0-/- mice) resemble cases of CMT1B and DSS with impaired myelin dosage (Martini et al., 1995a). Potential approaches to treat such diseases include the introduction of the normal gene in the nerves of strongly affected patients. In the present study we used P0-/- mice to evaluate the efficiency of a replication-defective, E1-deleted adenovirus vector carrying the lacZ (Ad-RSV-lacZ) or P0 (Ad-RSV-P0) gene to infect abnormally myelinating Schwann cells. The Ad-RSV-lacZ vector suspension was injected into the left sciatic nerve ofPO-/- mice and the nerves examined for beta-galactosidase activity by X-gal histochemistry. Contralateral nerves injected with vehicle solution or non-injected served as controls. Beta-galactosidase activity was detected in nerves injected with the Ad-RSV-lacZ vector up to 2 weeks post-injection. Immunosuppressing the mice with FK506 to decrease the infiltration of activated T-cells in infected nerves lengthened beta-galactosidase activity to 8 weeks, the longest time point examined. Ultrastructural analysis indicated that X-gal crystals were present mostly in abnormally myelinating Schwann cells. These findings demonstrate that an adenovirus vector can successfully infect Schwann cells in P0-/- mice and expression can be maintained for several weeks. The Ad-RSV-P0 suspension was then injected in the sciatic nerve of immunosuppressed P0-/- mice. Two and four weeks post-injection both P0 mRNA and protein could be detected by in situ hybridization and Western blotting in some of the nerves. Furthermore, P0 protein expression was observed in myelin-like structures and onion bulb-like cells by immunohistochemistry. These results indicate that Schwann cells in P0-/- mice can be induced to produce P0 protein after gene transfer. Genetic repair of abnormal Schwann cells by using adenovirus vectors might be a possible technique to treat animal models of inherited peripheral neuropathies.
 ...
PMID:Effective gene transfer of lacZ and P0 into Schwann cells of P0-deficient mice. 989 Jun 31

Myoblast transplantation (MT) may be a potential treatment for severe recessive hereditary myopathies. The limited results of MT in clinical trials led us to improve this technique in monkeys, an animal model phylogenetically similar to humans. Three Macaca mulata monkeys were used as donors and six as receivers for MT. Myoblasts were grown in culture from muscle biopsies of adult monkeys and infected with a retroviral vector encoding the LacZ gene. Different numbers of cells (i.e., 4 x 10(6), 8 x 10(6), and 24 x 10(6) cells) were transplanted into different muscles and 8 x 10(6) cells (resuspended in a notexin solution) were injected in one muscle of four monkeys. For these transplantations, the cell suspension (in a volume of about 100 microl) was injected at 35 sites less than 1 mm apart. Two other monkeys received 100 x 10(6) myoblasts resuspended in 1 ml of HBSS or 1 ml of notexin. For these two monkeys, the myoblasts were injected at 200-250 sites within a small portion of the muscle. All monkeys were immunosuppressed with daily injections of FK506. Four weeks after MT, the transplanted muscle portions were biopsied and the presence of beta-galactosidase-positive (beta-Gal+) muscle fibers was investigated. The number of beta-Gal+ fibers was 822 +/- 150 (site grafted with 4 x 10(6) cells), 1253 +/- 515 (8 x 10(6) cells), 1084 +/- 278 (24 x 10(6)), and 2852 +/- 1211 (notexin). In the monkeys grafted with 100 x 10(6) myoblasts, the number of beta-Gal+ fibers was 4850 (site without notexin) and 9600 (site with notexin). We demonstrated that a precise mechanical distribution of myoblasts into the tissue improves substantially MT in primates. The presence of notexin with the transplanted cells further increased the success of their transplantation. These are the best results obtained either with MT or gene therapy in primates and they encourage the possibility to human MT trials.
 ...
PMID:Successful myoblast transplantation in primates depends on appropriate cell delivery and induction of regeneration in the host muscle. 991 1

The steady-state level of the resident endoplasmic reticulum protein, 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR), is regulated, in part, by accelerated degradation in response to excess sterols or mevalonate. Previous studies of a chimeric protein (HM-Gal) composed of the membrane domain of HMGR fused to Escherichia coli beta-galactosidase, as a replacement of the normal HMGR cytosolic domain, have shown that the regulated degradation of this chimeric protein, HM-Gal, is identical to that of HMGR (Chun, K. T., Bar-Nun, S., and Simoni, R. D. (1990) J. Biol. Chem. 265, 22004-22010; Skalnik, D. G., Narita, H., Kent, C., and Simoni, R. D. (1988) J. Biol. Chem. 263, 6836-6841). Since the cytosolic domain can be replaced with beta-galactosidase without effect on regulated degradation, it has been assumed that the cytosolic domain was not important to this process and also that the membrane domain of HMGR was both necessary and sufficient for regulated degradation. In contrast to our previous results with HM-Gal, we observed in this study that replacement of the cytosolic domain of HMGR with various heterologous proteins can have an effect on the regulated degradation, and the effect correlates with the oligomeric state of the replacement cytosolic protein. Chimeric proteins that are oligomeric in structure are relatively stable, and those that are monomeric are unstable. To test the hypothesis that the oligomeric state of the cytosolic domain of HMGR influences degradation, we use an "inducible" system for altering the oligomeric state of a protein in vivo. Using a chimeric protein that contains the membrane domain of HMGR fused to three copies of FK506-binding protein 12, we were able to induce oligomerization by addition of a "double-headed" FK506-like "dimerizer" drug (AP1510) and to monitor the degradation rate of both the monomeric form and the drug-induced oligomeric form of the protein. We show that this chimeric protein, HM-3FKBP, is unstable in the monomeric state and is stabilized by AP1510-induced oligomerization. We also examined the degradation rate of HMGR as a function of concentrations within the cell. HMGR is a functional dimer; therefore, its oligomeric state and, we predict, its degradation rate should be concentration-dependent. We observed that it is degraded more rapidly at lower concentrations.
 ...
PMID:Oligomerization state influences the degradation rate of 3-hydroxy-3-methylglutaryl-CoA reductase. 1035 74

The goal of the present study was to determine the feasibility, success, and toxicity of myoblast transplantation (MT) in the whole muscle of primates. Allogenic myoblasts transduced with the beta-galactosidase (beta-Gal) gene were transplanted in the whole Biceps brachii of 5 monkeys immunosuppressed with FK506. Myoblast injections were spaced at every 1 to 1.5 mm in 7 muscles, as well as at every 5 mm in 2 muscles. Myoblasts were resuspended in HBSS, notexin 1 microg/ml or notexin 5 microg/ml. Depending on the number of beta-Gal labeled myoblasts and the injection protocol, biopsies of transplanted muscles exhibited 7% to 74% beta-Gal+ fibers 1 month after MT. Beta-Gal+ fibers were present in muscle biopsies made 3, 8, and 12 months after MT. Myoglobinuria and hyperkalemia, the risk factors after extensive muscle damage and notexin toxicity, were not observed. The withdrawal of immunosuppression led to histological evidences of cellular rejection of the graft. We concluded that MT can be successfully performed in large primate muscles without toxicity due to muscle damage. An effective immunosuppression allowed the maintenance of beta-Gal+ fibers up to 1 year after MT. These results suggest parameters that may allow effective MT in humans.
 ...
PMID:Myoblast transplantation in whole muscle of nonhuman primates. 1074 58

The upregulation of endogenous utrophin in skeletal muscle may lead to a new approach to the treatment of Duchenne muscular dystrophy (DMD). We found that injection of an E1, E3-deleted adenovirus vector expressing beta-galactosidase (beta-Gal) or green fluorescent protein (GFP) into the skeletal muscle of neonatal dystrophin-deficient mdx mice alleviated dystrophic pathology. In the adenovirus-infected muscles, an evaluation of sarcolemma stability showed low permeability and immunohistochemistry revealed utrophin upregulation at the extrasynaptic sarcolemma of mature muscle fibers. This utrophin upregulation was concomitant with endomysial cellular infiltration from a host immune reaction. There was no evidence of active muscle regeneration. In normal C57BL/10 mice, utrophin was also upregulated in adenovirus-injected skeletal muscles, where upregulated utrophin often coexisted with dystrophin. FK506 and anti-CD4 antibody administration decreased utrophin expression in adenovirus-injected mdx muscles and prevented the dystrophic phenotype from being mitigated, suggesting that an immune reaction is involved in utrophin upregulation. This is the first report demonstrating the improvement of the dystrophic phenotype as a result of the acquired overexpression of endogenous utrophin. Our findings provide an important clue to understanding the mechanism of utrophin expression and the development of an effective treatment for DMD.
 ...
PMID:Immune response to adenovirus-delivered antigens upregulates utrophin and results in mitigation of muscle pathology in mdx mice. 1075 47

The effect of pretreatments of host muscles with metalloproteinases (MMPs) or with notexin on the migration of transplanted myoblasts was investigated. Transgenic TnILacZ mice in which the beta-galactosidase gene is under the control of a quail fast skeletal troponin I gene promoter were used as donors. A polyethylene microtube with four perforations was inserted in the tibialis anterior (TA) of CD1 mice. Both pretreatment substances and cells were slowly injected through that microtube. Muscles were pretreated 2 days before myoblast injection either with a mixture of collagenase, matrilysin, and notexin or with only collagenase and matrilysin or only notexin. As control for our experiments, TnILacZ and C2C12 myoblasts were also injected in TA muscles not pretreated. Comparison of short and long-term myoblast radial migration was performed using a dye (PKH26) and X-gal staining, respectively. The recipient mice were immunosuppressed with FK506. Two days after myoblast transplantation, the cell movement in muscles pretreated with collagenase, matrilysin, and notexin was slightly greater than in muscles pretreated only with collagenase and matrilysin but was about twice that observed in muscles treated with notexin alone. Almost no radial migration of TnILacZ myoblasts was observed in untreated muscles. The C2C12 myoblasts showed a four-to fivefold higher migration capacity than TnILacZ myoblasts. At 15 days after TnILacZ myoblast transplantation, the farthest positive beta-gal muscle fibers show a two- to threefold extension of the initial migration observed at 2 days, demonstrating the ability of myoblasts to continue the migration following all pretreatments and even in the untreated muscles. In addition, more muscle fibers expressed the beta-gal reporter gene in muscles pretreated only with MMPs. Our results clearly demonstrate that muscle pretreatments with MMPs increase myoblast migration and fusion with host muscle fibers after transplantation and that the C2C12 cell line producing MMPs has a higher migratory capacity.
 ...
PMID:Intramuscular migration of myoblasts transplanted after muscle pretreatment with metalloproteinases. 1103 70

Limb girdle muscular dystrophy type 2B form and Miyoshi myopathy are both caused by mutations in the recently cloned gene dysferlin. In the present study, we have investigated whether cell transplantation could permit dysferlin expression in vivo. Two transplantation models were used: SCID mice transplanted with normal human myoblasts, and SJL mice, the mouse model for limb girdle muscular dystrophy type 2B and Miyoshi myopathy, transplanted with allogeneic primary mouse muscle cell cultures expressing the beta-galactosidase gene under control of a muscle promoter of Troponin I. FK506 immunosuppression was used in the non-compatible allogeneic model. One month after transplantation, human and mouse dysferlin proteins were detected in all transplanted SCID and SJL muscles, respectively. Co-localization of dysferlin and human dystrophin or beta-galactosidase-positive fibers was observed following the transplantation of myoblasts. Dysferlin proteins were monitored by immunocytochemistry and Western blot. The number of dysferlin-positive fibers was 40-50% and 20-30% in SCID and SJL muscle sections, respectively. Detection of dysferlin in both SCID mice and dysferlin-deficient SJL mouse shows that myoblast transplantation permits the expression of the donor dysferlin protein.
 ...
PMID:Dysferlin expression after normal myoblast transplantation in SCID and in SJL mice. 1173 59


<< Previous 1 2 3 Next >>