Fate and protective effect of marrow stromal cells after subretinal transplantation

Engraftment of marrow stromal cells (MSCs) has been proposed as a therapeutic approach for degenerative diseases. In this study we investigated the fate and dynamic progress of grafted MSCs in living retina with the aim of evaluating the use of transplanted MSCs to treat retinal degeneration. Approximately 1×10⁵ gfp -MSCs in 2 μl phosphate-buffered saline were injected into the subretinal space of adult Sprague-Dawley rats. Two weeks later, approximately 0.174%±0.082% of the transplanted cells had survived and diffused into the subretinal space. Nine weeks after transplantation the surviving gfp -MSCs accounted for 0.049%±0.023% of the number of cells injected and were mainly located at the injection site. The same number of MSCs were transplanted into the left eye subretinal space of 3-week-old hereditary retinal degenerative Royal College of Surgeons rats, and phosphate-buffered saline was injected into their right eyes as a control. Five weeks after transplantation, the amount of rudimentary photo-receptors was more significantly increased in grafted eyes than in control eyes. The results indicated that grafted MSCs could survive and rescue retinal degeneration.     

Phosphatized rare star-like mouth disc of Punctatus and its functional morphology from the earliest Cambrian of the South Shaanxi China

The Meishucun stage is the prelude in decipher-ing the Cambrian Explosion. In this prominent stage, rapid radioactive evolution and body-plan innovation have taken place and different associations of organism have been shaped. In this paper we report several 3D-preserved rare star-like fossils with finely preserved soft tissues which were recovered from the Kuanchuanpu Member of the Dengying Formation in South Shaanxi, China in 2003. By studying on functional morphology and analogy with mouthpart of Punctatus, there are evidences that this star-like organism approaches the coelenterates in systematic classification and the centre of star-like organism is its mouth. The appearance of coelenterates marks the real beginning of metazoan evolution. Therefore, it has the prominent position in the origin and evolutionary history of organisms. Perhaps the star-like organism represents the early types of coelenterate with original tentacles. These new materials provide new evidence for the origin, evolution and the functional evolution of the metazoan during the early stage of the Cambrian Explosion.     

Overexpression of PPK-1, the Caenorhabditis elegans Type I PIP kinase, inhibits growth cone collapse in the developing nervous system and causes axonal degeneration in adults

Growth cones are dynamic membrane structures that migrate to target tissue by rearranging their cytoskeleton in response to environmental cues. The lipid phosphatidylinositol (4,5) bisphosphate (PIP₂) resides on the plasma membrane of all eukaryotic cells and is thought to be required for actin cytoskeleton rearrangements. Thus PIP₂ is likely to play a role during neuron development, but this has never been tested in vivo. In this study, we have characterized the PIP₂ synthesizing enzyme Type I PIP kinase (ppk-1) in Caenorhabditis elegans. PPK-1 is strongly expressed in the nervous system, and can localize to the plasma membrane. We show that PPK-1 purified from C. elegans can generate PIP₂in vitro and that overexpression of the kinase causes an increase in PIP₂ levels in vivo. In developing neurons, PPK-1 overexpression leads to growth cones that become stalled, produce ectopic membrane projections, and branched axons. Once neurons are established, PPK-1 overexpression results in progressive membrane overgrowth and degeneration during adulthood. These data suggest that overexpression of the Type I PIP kinase inhibits growth cone collapse, and that regulation of PIP₂ levels in established neurons may be important to maintain structural integrity and prevent neuronal degeneration.     

DIF-1 induces the basal disc of the Dictyostelium fruiting body

The polyketide DIF-1 induces Dictyostelium amoebae to form stalk cells in culture. To better define its role in normal development, we examined the phenotype of a mutant blocking the first step of DIF-1 synthesis, which lacks both DIF-1 and its biosynthetic intermediate, dM-DIF-1 (des-methyl-DIF-1). Slugs of this polyketide synthase mutant (stlB⁻) are long and thin and rapidly break up, leaving an immotile prespore mass. They have ∼ 30% fewer prestalk cells than their wild-type parent and lack a subset of anterior-like cells, which later form the outer basal disc. This structure is missing from the fruiting body, which perhaps in consequence initiates culmination along the substratum. The lower cup is rudimentary at best and the spore mass, lacking support, slips down the stalk. The dmtA⁻ methyltransferase mutant, blocked in the last step of DIF-1 synthesis, resembles the stlB⁻ mutant but has delayed tip formation and fewer prestalk-O cells. This difference may be due to accumulation of dM-DIF-1 in the dmtA⁻ mutant, since dM-DIF-1 inhibits prestalk-O differentiation. Thus, DIF-1 is required for slug migration and specifies the anterior-like cells forming the basal disc and much of the lower cup; significantly the DIF-1 biosynthetic pathway may supply a second signal - dM-DIF-1.     

Growth factor effects on costal chondrocytes for tissue engineering fibrocartilage

Tissue-engineered fibrocartilage could become a feasible option for replacing tissues such as the knee meniscus or temporomandibular joint disc. This study employed five growth factors (insulin-like growth factor-I, transforming growth factor-beta1, epidermal growth factor, platelet-derived growth factor-BB, and basic fibroblast growth factor) in a scaffoldless approach with costal chondrocytes, attempting to improve biochemical and mechanical properties of engineered constructs. Samples were quantitatively assessed for total collagen, glycosaminoglycans, collagen type I, collagen type II, cells, compressive properties, and tensile properties at two time points. Most treated constructs had lower biomechanical and biochemical properties than the controls with no growth factors, suggesting a detrimental effect, but the treatment with insulin-like growth factor-I tended to improve the constructs. Additionally, the 6-week time point was consistently better than that at 3 weeks, with total collagen, glycosaminoglycans, and aggregate modulus doubling during this time. Further optimization of the time in culture and exogenous stimuli will be important in making a more functional replacement tissue.     

Shox2-deficiency leads to dysplasia and ankylosis of the temporomandibular joint in mice

The temporomandibular joint (TMJ) is a unique synovial joint whose development differs from the formation of other synovial joints. Mutations have been associated with the developmental defects of the TMJ only in a few genes. In this study, we report the expression of the homeobox gene Shox2 in the cranial neural crest derived mesenchymal cells of the maxilla-mandibular junction and later in the progenitor cells and undifferentiated chondrocytes of the condyle as well as the glenoid fossa of the developing TMJ. A conditional inactivation of Shox2 in the cranial neural crest-derived cells causes developmental abnormalities in the TMJ, including dysplasia of the condyle and glenoid fossa. The articulating disc forms but fuses with the fibrous layers of the condyle and glenoid fossa, clinically known as TMJ ankylosis. Histological examination indicates a delay in development in the mutant TMJ, accompanied by a significantly reduced rate of cell proliferation. In situ hybridization further demonstrates an altered expression of several key osteogenic genes and a delayed expression of the osteogenic differentiation markers. Shox2 appears to regulate the expression of osteogenic genes and is essential for the development and function of the TMJ. The Shox2 conditional mutant thus provides a unique animal model of TMJ ankylosis.     

Immunolocalization of MMP-19 in the human intervertebral disc: implications for disc aging and degeneration

Matrix metalloproteinases (MMPs) degrade components of the extracellular matrix of the disc, but the presence of MMP-19 has not been explored. In other tissues, MMP-19 is known to act in proteolysis of the insulin-like growth factor (IGF) binding protein-3, thereby exposing this protein to make it available to influence cell behavior. MMP-19 also has been shown to inhibit capillary-like formation and thus play a role in the avascular nature of the disc. Using immunohistochemistry, normal discs from six subjects aged newborn through 10 years and 20 disc specimens from control donors or surgical patients aged 15-76 (mean age 40.2 years) were examined for immunolocalization of MMP-19; six Thompson grade I discs, five Thompson grade II, eight Thompson grade III, five Thompson grade IV, and one Thompson grade V discs were analyzed. The results indicate that in discs from young subjects, MMP-19 was uniformly localized in the outer annulus. In discs from adult donors and surgical patients, outer and inner annulus cells only occasionally showed MMP-19 localization. The greatest expression of MMP-19 was observed in young discs, and little expression was seen in older or degenerating discs. Because MMP-19 has been shown to regulate IGF-mediated proliferation in other tissues, its decline in the aging/degenerating disc may contribute to the age-related decrease in disc cell numbers.     

Retinal isoforms of inosine 5'-monophosphate dehydrogenase type 1 are poor nucleic acid binding proteins

The RP 10 form of autosomal dominant retinitis pigmentosa (adRP) is caused by mutations in the widely expressed protein inosine 5′-monophosphate dehydrogenase type 1 (IMPDH1). These mutations have no effect on the enzymatic activity of IMPDH1, but do perturb the association of IMPDH1 with nucleic acids. Two newly discovered retinal-specific isoforms, IMPDH1(546) and IMPDH1(595), may provide the key to the photoreceptor specificity of disease [S.J. Bowne, Q. Liu, L.S. Sullivan, J. Zhu, C.J. Spellicy, C.B. Rickman, E.A. Pierce, S.P. Daiger, Invest. Ophthalmol. Vis. Sci. 47 (2006) 3754-3765]. Here we express and characterize the normal IMPDH1(546) and IMPDH1(595), together with their adRP-linked variants, D226N. The enzymatic activity of the purified IMPDH1(546), IMPDH1(595) and the D226N variants is indistinguishable from the canonical form. The intracellular distribution of IMPDH1(546) and IMPDH1(595) is also similar to the canonical IMPDH1 and unaffected by the D226N mutation. However, unlike the canonical IMPDH1, the retinal specific isoforms do not bind significant fractions of a random pool of oligonucleotides. This observation indicates that the C-terminal extension unique to the retinal isoforms blocks the nucleic acid binding site of IMPDH1, and thus uniquely regulates protein function within photoreceptors.     

Mechanistic Studies of ABCR, the ABC Transporter in Photoreceptor Outer Segments Responsible for Autosomal Recessive Stargardt Disease

ABCR is an ABC transporter that is found exclusively in vertebrate photoreceptor outer segments. Mutations in the human ABCR gene are responsible for autosomal recessive Stargardt disease, the most common cause of early onset macular degeneration. In this paper we review our recent work with purified and reconstituted ABCR derived from bovine retina and from cultured cells expressing wild type or site-directed mutants of human ABCR. These experiments implicate all-trans-retinal (or Schiff base adducts between all-trans-retinal and phosphatidylethanolamine) as the transport substrate, and they reveal asymmetric roles for the two nucleotide binding domains in the transport reaction. A model for the retinal transport reaction is presented which accounts for these experimental observations.