Loss of nuclear poly(A)-binding protein 1 causes defects in myogenesis and mRNA biogenesis

The nuclear poly(A)-binding protein 1 (PABPN1) is a ubiquitously expressed protein that plays a critical role in polyadenylation. Short expansions of the polyalanine tract in the N-terminus of PABPN1 lead to oculopharyngeal muscular dystrophy (OPMD), which is an adult onset disease characterized by eyelid drooping, difficulty in swallowing and weakness in the proximal limb muscles. Although significant data from in vitro biochemical assays define the function of PABPN1 in control of poly(A) tail length, little is known about the role of PABPN1 in mammalian cells. To assess the function of PABPN1 in mammalian cells and specifically in cells affected in OPMD, we examined the effects of PABPN1 depletion using siRNA in primary mouse myoblasts from extraocular, pharyngeal and limb muscles. PABPN1 knockdown significantly decreased cell proliferation and myoblast differentiation during myogenesis in vitro. At the molecular level, PABPN1 depletion in myoblasts led to a shortening of mRNA poly(A) tails, demonstrating the cellular function of PABPN1 in polyadenylation control in a mammalian cell. In addition, PABPN1 depletion caused nuclear accumulation of poly(A) RNA, revealing that PABPN1 is required for proper poly(A) RNA export from the nucleus. Together, these experiments demonstrate that PABPN1 plays an essential role in myoblast proliferation and differentiation, suggesting that it is required for muscle regeneration and maintenance in vivo.

Pkc1 acts through Zds1 and Gic1 to suppress growth and cell polarity defects of a yeast eIF5A mutant

eIF5A is a highly conserved putative eukaryotic translation initiation factor that has been implicated in translation initiation, nucleocytoplasmic transport, mRNA decay, and cell proliferation, but with no precise function assigned so far. We have previously shown that high-copy PKCI suppresses the phenotype of tif51A-1, a temperature-sensitive mutant of eIF5A in S. cerevisiae. Here, in an attempt to further understand how Pkc1 functionally interacts with eIF-5A, it was determined that PKCI suppression of tif51A-1 is independent of the cell integrity MAP kinase cascade. Furthermore, two new suppressor genes, ZDS1 and GIC1, were identified. We demonstrated that ZDS1 and ZDS2 are necessary for PKC1, but not for GIC1 suppression. Moreover, high-copy GIC1 also suppresses the growth defect of a PKCI mutant (stt1), suggesting the existence of a Pkc1-Zds1-Gic1 pathway. Consistent with the function of Gic1 in actin organization, the tif51A-1 strain shows an actin polarity defect that is partially recovered by overexpression of Pkc1 and Zds1 as well as Gic1. Additionally, PCL1 and BNI1, important regulators of yeast cell polarity, also suppress tif51A-1 temperature sensitiviiy Taken together, these data strongly Support the correlated involvement of Pkc1 and eIF5A in establishing actin polarity, which is essential for bud formation and G1/S transition in S. cerevisiae.

Toward an understanding of intermediate- and short-range defects in ZnO single crystals. A combined experimental and theoretical study

A joint use of experimental and theoretical techniques allows us to understand the key role of intermediate- and short-range defects in the structural and electronic properties of ZnO single crystals obtained by means of both conventional hydrothermal and microwave-hydrothermal synthesis methods. X-ray diffraction, Raman spectra, photoluminescence, scanning electronic and transmission electron microscopies were used to characterize the thermal properties, crystalline and optical features of the obtained nano and microwires ZnO structures. In addition, these properties were further investigated by means of two periodic models, crystalline and disordered ZnO wurtzite structure, and first principles calculations based on density functional theory at the B3LYP level. The theoretical results indicate that the key factor controlling the electronic behavior can be associated with a symmetry breaking process, creating localized electronic levels above the valence band.

Treatment of segmental tibial defects using acute bone shortening followed by gradual lengthening with circular external fixator

The aim of this study was to clinically and radiographically evaluate acute bone shortening followed by gradual lengthening in the treatment of large segmental tibia defects induced in seven clinically normal dogs. A circular external fixator was assembled with one proximal 5/8-circle ring, one middle ring and one distal ring connected with three rods. Thirty per cent of the tibia and fibula were removed in the middle and distal parts of the diaphyses, between the middle and distal rings. Acute bone shortening with compression of proximal and distal segments was performed. A subperiosteal osteotomy was performed between the half-ring and middle ring. Bone distraction started 7 days after surgery; after lengthening, the apparatus was left in place for 14 weeks for consolidation of regenerated bone. The frame was removed at the end of this period, and the dogs observed for four more weeks. Functional results were considered excellent in two, good in three and fair in the other two dogs. Bone regeneration within the distraction gap was obtained 14 weeks after neutral fixation period. We concluded that acute bone shortening followed by gradual lengthening by Ilizarov method can be used to treat extensive tibial defects in dogs, although it presents limb temporary abnormal limb shape and unequal length as early disadvantages.

Vegetal polymer in repair of defects of the orbital floor: an experimental study in rabbits

Background: An experimental study was done to assess the ability of the vegetal polymer miniplates and screws to repair defects of the orbital floor.Methods: An artificial standard-sized defect was created in the bony floor of right orbit of 45 albino rabbits. The animals were divided into three experimental groups: control group (G1) involving animals with orbital floor defect and no treatment; titanium group (G2) containing animals with orbital floor defect repaired by titanium miniplates and screws; vegetal polymer group (G3) composed of animals with similar orbital floor defects repaired by vegetal polymer miniplates and screws. Throughout the course of the experiment, the animals were clinically evaluated. At 15, 30 and 60 days after surgery, the animals were killed. They were X-rayed immediately after the floor defect and at the moment of sacrifice. Histological and morphometric evaluation of inflammatory reaction and bone healing was done. Data were statistically evaluated.Results: No implants were extruded. Bone consolidation was similar in G2 and G3 and better than in G1 group animals. Inflammatory reaction was most pronounced in animals of G3 15 days after surgery, and it subsided over time.Conclusion: Vegetal polymer miniplates and screws induces small inflammatory reaction and had the ability to stimulate bone growth with good integration in the orbital floor defect allowing to consider the vegetal polymer adequate option to treat orbital floor defects. Future studies involving long-term follow-up and biomechanical tests to evaluate material resistance to traction are needed.

Miniplate fixation for the repair of segmental mandibular defects filled with autogenous bone in cats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Influence of the ratio of particulate autogenous bone graft/platelet-rich plasma on bone healing in critical-size defects: a histologic and histometric study in rat calvaria

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Bone healing in critical-size defects treated with platelet-rich plasma activated by two different methods. A histologic and histometric study in rat calvaria

The purpose of this study was to analyze histologically the influence of platelet-rich plasma (PRP) coagulated with two different activators on bone healing in surgically created critical-size defects (CSD) in rat calvaria.Forty-eight rats were divided into three groups: C, PRP-C and PRP-T. An 8 mm diameter CSD was created in the calvarium of each animal. In group C, the defect was filled by a blood clot only. In groups PRP-C and PRP-T, the defect was filled with PRP activated with either calcium chloride or thromboplastin solution, respectively. Each group was divided into two subgroups (n = 8 per subgroup) and killed at either 4 or 12 weeks postoperatively. Histologic and histometric analyses were performed. The amount of new bone formed was calculated as a percentage of the total area of the original defect. Percentage data were transformed into arccosine for statistical analysis (analysis of variance, Tukey's post hoc test, p < 0.05).No defect completely regenerated with bone. Group PRP-C had a statistically greater amount of bone formation than groups C and PRP-T at both time points of analysis. No statistically significant differences were observed between groups C and PRP-T.It can be concluded that the type of activator used to initiate PRP clot formation influences its biological effect on bone healing in CSD in rat calvaria.

Effect of alveolex on the bone defects repair stimulated by rhBMP-2: Histomorphometric study

OBJECTIVE: The aim of this study was to evaluate histomorphometrically the effect of alveolex (Propolis 10%) on the repair of bone cavities in the calvaria of rats. MATERIALS and METHODS: A 5 mm diameter bone defect was made in the calvaria of male Wistar rats using the drill-type trephine. The defects were filled with rhBMP-21Alveolex, rhBMP-2, Alveolex, or coagulum. Twenty-eight animals with seven subjects on each were sacrificed 30 days after surgery and samples were fixed and embedded in paraffin. Histological sections stained by HE (hematoxylin and eosin) were obtained from the calvaria bone defect and analyzed by a differential point-counting method. RESULTS: Group I and II, rhBMP-21Alveolex and rhBMP-2, respectively, presented higher levels of newly formed bone than other groups (P < 0.001). There were not significant differences between groups I and II (P > 0.05). In addition, there was not significant difference between groups III and IV, Control-Coagulum and Alveolex, respectively (P > 0.05). CONCLUSION: Alveolex has increased the bone repair in calvaria defects of rats when associated to rhBMP-2, however without significant differences for rhBMP-2 isolated group; Alveolex isolated group showed the lowest levels of newly formed bone with no significant differences to coagulum group (control). Microsc. Res. Tech. 75: 36-41, 2012. (C) 2011 Wiley Periodicals, Inc.

The effect of BMP-2 on the osteoconductive properties of beta-tricalcium phosphate in rat calvaria defects

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Osteoconductive Properties of beta-Tricalcium Phosphate Matrix, Polylactic and Polyglycolic Acid Gel, and Calcium Phosphate Cement in Bone Defects

Extensive bone defects in maxillofacial region can be corrected with autogenous grafts; otherwise, the disadvantages of the therapeutics modality take the research for new bone substitutes. The aim of the study was to evaluate and compare the osteoconductive properties of 3 commercial available biomaterials. A total of 30 calvarial defects (5-mm diameter) were randomly divided into 5 treatment groups, with a total of 6 defects per treatment group (n = 6). The treatment groups were as follows: 500 to 1000 Km beta-tricalcium phosphate (beta-TCP), polylactic and polyglycolic acid (PL/PG) gel, calcium phosphate cement, untreated control, and autograft control. The evaluations were based on histomorphometric analysis at 60 postoperative days. The results have shown that beta-TCP and autograft control supported bone formation at 60 postoperative days. beta-Tricalcium phosphate showed the highest amount of mineralized area per total area and statistically significant compared with PL/PG, calcium phosphate cement, and untreated control groups. The PL/PG gel does not have osteoconductive properties and performed similar to empty control. Calcium phosphate cement showed higher number of multinucleated giant cells around the sites of the biomaterial and showed newly formed bone only at the edges of the biomaterial, without bone formation within the biomaterial. The findings presented herein indicate that bone formation reached a maximum level when rat calvarial defects were filled with beta-TCP at 60 postoperative days. Further studies should be conducted with beta-TCP to understand the potential of this biomaterial in bone regeneration.

Effect of platelet-rich plasma on bone healing of autogenous bone grafts in critical-size defects

AimThis study histologically analysed the effect of autogenous platelet-rich plasma (PRP), prepared according to a new semiautomatic system, on healing of autogenous bone (AB) grafts placed in surgically created critical-size defects (CSD) in rabbit calvaria.Material and MethodsSixty rabbits were divided into three groups: C, AB and AB/PRP. A CSD was created in the calvarium of each animal. In Group C (control), the defect was filled by blood clot only. In Group AB (autogenous bone graft), the defect was filled with particulate autogenous bone. In Group AB/PRP (autogenous bone graft with platelet-rich plasma), it was filled with particulate autogenous bone combined with PRP. All groups were divided into subgroups (n=10) and euthanized at 4 or 12 weeks post-operatively. Histometric and histologic analyses were performed. Data were statistically analysed (anova, t-test, p < 0.05).ResultsGroup C presented significantly less bone formation compared with Group AB and AB/PRP in both periods of analysis (p < 0.001). At 4 weeks, Group AB/PRP showed a statistically greater amount of bone formation than Group AB (64.44 +/- 15.0% versus 46.88 +/- 14.15%; p=0.0181). At 12 weeks, no statistically significant differences were observed between Groups AB and AB/PRP (75.0 +/- 8.11% versus 77.90 +/- 8.13%; p > 0.05). It is notable that the amount of new bone formation in Group AB/PRP at 4 weeks was similar to that of Group AB at 12 weeks (p > 0.05).ConclusionWithin its limitation, the present study has indicated that (i) AB and AB/PRP significantly improved bone formation and (ii) a beneficial effect of PRP was limited to an initial healing period of 4 weeks.

Influence of Bioactive Glass and/or Acellular Dermal Matrix on Bone Healing of Surgically Created Defects in Rat Tibiae: A Histological and Histometric Study

The purpose of this study was to histologically analyze the influence of bioactive glass and/or acellular dermal matrix on bone healing in surgically created defects in the tibiae of 64 rats (Rattus norvegicus, albinus, Wistar). Materials and Methods: A 4-mm X 3-mm unicortical defect was created on the anterolateral surface of the tibia. Animals were divided into 4 groups: C, control; BG, the defect was filled with bioactive glass; ADM, the defect was covered with acellular dermal matrix; and BG/ADM, the defect was filled with bioactive glass and covered with acellular dermal matrix. Animals were sacrificed at 10 or 30 days postoperatively, and the specimens were removed for histologic processing. The formation of new bone in the cortical area of the defect was evaluated histomorphometrically. Results: At 10 and 30 days postoperatively, groups C (39.65% +/- 5.63% / 63.34% +/- 5.22%) and ADM (38.12% +/- 5.53 / 58.96% +/- 7.05%) presented a larger amount of bone formation compared to the other groups (P<.05). In the same periods, groups BG (13.10% +/- 6.29% / 29.5% +/- 5.56%) and BG/ADM (20.72% +/- 8.31% / 24.19% +/- 6.69%) exhibited statistically similar new bone formation. However, unlike the other groups, group BG/ADM did not present a significant increase in bone formation between the 2 time points. Conclusion: Based on these results, it can be concluded that all of the materials used in this study delayed bone healing in non-critical-size defects. INT J ORAL MAXILLOFAC IMPLANTS 2008;23:811-817

Use of Titanium Mesh for Reconstruction of Extensive Defects in Fronto-orbito-ethmoidal Fracture

Traumatic injuries treatment of the fronto-naso-orbito-ethmoidal region has been one of the most challenging treatments within maxillofacial surgery, particularly of extensive orbital defects, very common in this type of pathologic condition. A 48-year-old man involved in a car collision presented an extensive bilateral fracture of the orbit medial wall, nasal bones, the nasal septum, and the frontal anterior table. The clinical and tomographic findings concluded the diagnosis of a maxilla and fronto-naso-orbito-ethmoidal fracture. Among the variety of biomaterials, the titanium mesh was elected because of the extension and magnitude of the bone defect, obtaining this way esthetic and functional results with better prognosis.

Repair process of surgical defects filled with autogenous bone grafts in tibiae of diabetic rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)