Optimization of alginate microcapsules containing cells overexpressing α-l-iduronidase using Box-Behnken design.

Mucopolysaccharidosis type I (MPS I) is an autosomal recessive disease caused by deficiency of α-l-iduronidase (IDUA), which results in the lysosomal accumulation of glycosaminoglycans (GAG) leading to widespread clinical manifestations. The microencapsulation of IDUA overexpressing recombinant cells has been considered as a promising strategy for the treatment of MPS I. This study aimed at the optimization of alginate microcapsules containing recombinant BHK (Baby Hamster Kidney) cells (rBHK) overexpressing IDUA produced by electrostatic extrusion technique.

Subcutaneous implantation of microencapsulated cells overexpressing α-L-iduronidase for mucopolysaccharidosis type I treatment.

Mucopolysaccharidosis type I (MPS I) is caused by a deficiency of α-L-iduronidase (IDUA), resulting in accumulation of glycosaminoglycans (GAG) in lysosomes. Microencapsulation of recombinant cells is a promising gene/cell therapy approach that could overcome the limitations of the current available treatments. In the present study we produced alginate-poly-L-lysine-alginate (APA) microcapsules containing recombinant cells overexpressing IDUA, which were implanted in the subcutaneous space of MPS I mice in order to evaluate their potential effect as a treatment for this disease.

Experimental evidence that bioenergetics disruption is not mainly involved in the brain injury of glutaryl-CoA dehydrogenase deficient mice submitted to lysine overload.

Bioenergetics dysfunction has been postulated as an important pathomechanism of brain damage in glutaric aciduria type I, but this is still under debate. We investigated activities of citric acid cycle (CAC) enzymes, lactate release, respiration and membrane potential (ΔΨm) in mitochondrial preparations from cerebral cortex and striatum of 30-day-old glutaryl-CoA dehydrogenase deficient (Gcdh-/-) and wild type mice fed a baseline or a high lysine (Lys, 4.7%) chow for 60 or 96h.

Increased glutamate receptor and transporter expression in the cerebral cortex and striatum of gcdh-/- mice: possible implications for the neuropathology of glutaric acidemia type I.

We determined mRNA expression of the ionotropic glutamate receptors NMDA (NR1, NR2A and NR2B subunits), AMPA (GluR2 subunit) and kainate (GluR6 subunit), as well as of the glutamate transporters GLAST and GLT1 in cerebral cortex and striatum of wild type (WT) and glutaryl-CoA dehydrogenase deficient (Gchh-/-) mice aged 7, 30 and 60 days. The protein expression levels of some of these membrane proteins were also measured. Overexpression of NR2A and NR2B in striatum and of GluR2 and GluR6 in cerebral cortex was observed in 7-day-old Gcdh-/-.

In vivo experimental evidence that the major metabolites accumulating in 3-hydroxy-3-methylglutaryl-CoA lyase deficiency induce oxidative stress in striatum of developing rats: a potential pathophysiological mechanism of striatal damage in this disorder.

3-Hydroxy-3-methylglutaryl-CoA lyase (HL) deficiency is a genetic disorder biochemically characterized by predominant accumulation of 3-hydroxy-3-methylglutaric (HMG) and 3-methylglutaric (MGA) acids in tissues and biological fluids of affected individuals. Clinically, the patients present neurological symptoms and basal ganglia injury, whose pathomechanisms are partially understood. In the present study, we investigated the ex vivo effects of intrastriatal administration of HMG and MGA on important parameters of oxidative stress in striatum of developing rats.

Treatment of MPS I mice with microencapsulated cells overexpressing IDUA: effect of the prednisolone administration.

Cell encapsulation, although a promising strategy to deliver therapeutic products, is hampered by immune response against biomaterials. The aim of this article is to assess the effect of prednisolone on enzyme release by microencapsulated cells implanted in vivo. Recombinant cells encapsulated were implanted in the peritoneum of wild-type mice and mucopolysaccharidosis (MPS) I mice, with or without prednisolone.

Chloramphenicol enhances IDUA activity on fibroblasts from mucopolysaccharidosis I patients.

Mucopolysaccharidosis I (MPS I) is a genetic disorder caused by mutations on α-L-iduronidase (IDUA) gene, leading to low or null enzyme activity. As nonsense mutations are present in about two thirds of the patients, stop codon read through (SCRT) is a potential alternative to achieve enhanced enzyme activity. This mechanism suppresses premature stop codon mutations allowing the protein to be fully translated.

Influence of phospholipid composition on cationic emulsions/DNA complexes: physicochemical properties, cytotoxicity, and transfection on Hep G2 cells.

Background: Cationic nanoemulsions have been recently considered as potential delivery systems for nucleic acids. This study reports the influence of phospholipids on the properties of cationic nanoemulsions/DNA plasmid complexes.

Methods: Nanoemulsions composed of medium-chain triglycerides, stearylamine, egg lecithin or isolated phospholipids, ie, DSPC, DOPC, DSPE, or DOPE, glycerol, and water were prepared by spontaneous emulsification.

Cell microencapsulation: a potential tool for the treatment of neuronopathic lysosomal storage diseases.

Lysosomal storage disorders (LSD) are monogenic diseases caused by the deficiency of different lysosomal enzymes that degrade complex substrates such as glycosaminoglycans, sphingolipids, and others. As a consequence there is multisystemic storage of these substrates. Most treatments for these disorders are based in the fact that most of these enzymes are soluble and can be internalized by adjacent cells via mannose-6-phosphate receptor.

Effects of cryopreservation and hypothermic storage on cell viability and enzyme activity in recombinant encapsulated cells overexpressing alpha-L-iduronidase.

Here, we show the effects of cryopreservation and hypothermic storage upon cell viability and enzyme release in alginate beads containing baby hamster kidney cells overexpressing alpha-L-iduronidase (IDUA), the enzyme deficient in mucopolysaccharidosis type I. In addition, we compared two different concentrations of alginate gel (1% and 1.5%) in respect to enzyme release from the beads and their shape and integrity.

Clinical and biochemical study of 29 Brazilian patients with metachromatic leukodystrophy.

Metachromatic leukodystrophy (MLD) is a lysosomal disorder caused by arylsulfatase A (ARSA) deficiency. It is classified into three forms according to the age of onset of symptoms (late infantile, juvenile, and adult). We carried out a cross-sectional and retrospective study, which aimed to determine the epidemiological, clinical, and biochemical profile of MLD patients from a national reference center for Inborn Errors of Metabolism in Brazil.

In vitro correction of ARSA deficiency in human skin fibroblasts from metachromatic leukodystrophy patients after treatment with microencapsulated recombinant cells.

Metachromatic leukodystrophy (MLD) is an autosomal recessive disorder due to arylsulfatase A (ARSA) deficiency that affects primarily the central nervous system. Ongoing treatments include enzyme replacement therapy and bone marrow transplantation, both limited in their effects due to the blood-brain barrier. An alternative approach would be the in situ implantation of encapsulated cells over expressing ARSA.

B subunit of Escherichia coli heat-labile enterotoxin as adjuvant of humoral immune response in recombinant BCG vaccination.

The B subunit of Escherichia coli heat-labile enterotoxin (LTB), a nontoxic molecule with potent biological properties, is a powerful mucosal and parenteral adjuvant that induces a strong immune response against co-administered or coupled antigens. In this paper, the effect of LTB on the humoral immune response to recombinant BCG (rBCG) vaccination was evaluated. Isogenic mice were immunized with rBCG expressing the R1 repeat region of the P97 adhesin of Mycoplasma hyopneumoniae alone (rBCG/R1) or fused to LTB (rBCG/LTBR1).