Properties of neurons derived from induced pluripotent stem cells of Gaucher disease type 2 patient fibroblasts: potential role in neuropathology.

Gaucher disease (GD) is caused by insufficient activity of acid β-glucosidase (GCase) resulting from mutations in GBA1. To understand the pathogenesis of the neuronopathic GD, induced pluripotent stem cells (iPSCs) were generated from fibroblasts isolated from three GD type 2 (GD2) and 2 unaffected (normal and GD carrier) individuals. The iPSCs were converted to neural precursor cells (NPCs) which were further differentiated into neurons.

Collagen V is a dominant regulator of collagen fibrillogenesis: dysfunctional regulation of structure and function in a corneal-stroma-specific Col5a1-null mouse model.

Collagen V is a regulatory fibril-forming collagen that forms heterotypic fibrils with collagen I. Deletion of collagen V in the mouse is associated with a lack of fibril assembly in the embryonic mesenchyme, with a resultant lethal phenotype. The current work elucidates the regulatory roles of collagen V during development and growth of tissues.

Regulation of collagen fibril nucleation and initial fibril assembly involves coordinate interactions with collagens V and XI in developing tendon.

Collagens V and XI comprise a single regulatory type of fibril-forming collagen with multiple isoforms. Both co-assemble with collagen I or II to form heterotypic fibrils and have been implicated in regulation of fibril assembly. The objective of this study was to determine the roles of collagens V and XI in the regulation of tendon fibrillogenesis.

Three-dimensional in vitro effects of compression and time in culture on aggregate modulus and on gene expression and protein content of collagen type II in murine chondrocytes.

The objectives of this study were to determine how culture time and dynamic compression, applied to murine chondrocyte-agarose constructs, influence construct stiffness, expression of col2 and type II collagen. Chondrocytes were harvested from the ribs of six newborn double transgenic mice carrying transgenes that use enhanced cyan fluorescent protein (ECFP) and green fluorescent protein (GFP-T) as reporters for expression from the col2a1 and col1a1 promoters, respectively. Sixty-three constructs (8 mm diameter x 3 mm thick) per animal were created by seeding chondrocytes (10 x 10(6) per mL) in agarose gel (2% w/v).

ColVa1 and ColXIa1 are required for myocardial morphogenesis and heart valve development.

Genetic mutations in minor fibrillar collagen types Va1 (ColVa1) and XIa1 (ColXI) have been identified in connective tissue disorders including Ehlers-Danlos syndrome and chondrodysplasias. ColVa1+/- and ColXIa1-/- mutant mice recapitulate these human disorders and show aberrations in collagen fiber organization in connective tissue of the skin, cornea, cartilage, and tendon. In the heart, fibrous networks of collagen fibers form throughout the ventricular myocardium and heart valves, and alterations in collagen fiber homeostasis are apparent in many forms of cardiac disease associated with myocardial dysfunction and valvular insufficiency.

Murine model of the Ehlers-Danlos syndrome. col5a1 haploinsufficiency disrupts collagen fibril assembly at multiple stages.

The most commonly identified mutations causing Ehlers-Danlos syndrome (EDS) classic type result in haploinsufficiency of proalpha1(V) chains of type V collagen, a quantitatively minor collagen that co-assembles with type I collagen as heterotypic fibrils. To determine the role(s) of type I/V collagen interactions in fibrillogenesis and elucidate the mechanism whereby half-reduction of type V collagen causes abnormal connective tissue biogenesis observed in EDS, we analyzed mice heterozygous for a targeted inactivating mutation in col5a1 that caused 50% reduction in col5a1 mRNA and collagen V. Comparable with EDS patients, they had decreased aortic stiffness and tensile strength and hyperextensible skin with decreased tensile strength of both normal and wounded skin.

Endogenously expressed multimeric self-cleaving hammerhead ribozymes ablate mutant collagen in cellulo.

Hammerhead ribozymes are small catalytic RNA molecules that can be targeted to any RNA molecule containing a putative cleavage site. We developed a vector (pCOLZ) that uses the COL1A1 promoter to drive expression of a self-cleaving multimeric ribozyme (M8Rz547) and its monomeric counterpart (Rz547). The ribozymes were stably coexpressed in MC3T3-E1 osteoblasts expressing a truncated COL1A1 target transcript.

Type V collagen controls the initiation of collagen fibril assembly.

Vertebrate collagen fibrils are heterotypically composed of a quantitatively major and minor fibril collagen. In non-cartilaginous tissues, type I collagen accounts for the majority of the collagen mass, and collagen type V, the functions of which are poorly understood, is a minor component. Type V collagen has been implicated in the regulation of fibril diameter, and we reported recently preliminary evidence that type V collagen is required for collagen fibril nucleation (Wenstrup, R.

Reduced type I collagen utilization: a pathogenic mechanism in COL5A1 haplo-insufficient Ehlers-Danlos syndrome.

To examine mechanisms by which reduced type V collagen causes weakened connective tissues in the Ehlers-Danlos syndrome (EDS), we examined matrix deposition and collagen fibril morphology in long-term dermal fibroblast cultures. EDS cells with COL5A1 haplo-insufficiency deposited less than one-half of hydroxyproline as collagen compared to control fibroblasts, though total collagen synthesis rates are near-normal because type V collagen represents a small fraction of collagen synthesized. Cells from patients with osteogenesis imperfecta (OI) and haplo-insufficiency for proalpha1(I) chains of type I collagen also incorporated about one-half the collagen as controls, but this amount was proportional to their reduced rates of total collagen synthesis.