Journal of Tissue Science and Engineering

In order to establish the convertibility of a host for bone augmentation, we herein developed a new honeycombshaped β-tricalcium phosphate (37H) using atelocollagen as a scaffold, which exhibited unique geometric properties for in vitro and in vivo studies.
Human mesenchymal stem cells (MSC) were cultured with 37H or atelocollagen-coated honeycomb-shaped β-tricalcium phosphate (Col37H), and their osteoblastic differentiation was then analyzed. Atelocollagen promoted cell adhesion and formation of vessel-like structures in the tunnels of scaffolds of cultured MSC. The mRNA expression levels of type I collagen, osteocalcin, and VEGF were greater in MSC cultured with Col37H than with 37H. Bone generation with Col37H in the rat calvaria was greater than with 37H, and this was attributed to early vessel construction. A large number of blood vessels invaded tunnels from the periosteum and existing bone surface. A strong VEGF signal was detected immediately before the new bone surface in the tunnels of Col37H.
These results indicate that the addition of atelocollagen to Col37H has potential in the construction of functional artificial bone.

Endothelial cells (EC) have important physiological functions, and they may also have a role in pathology. To better understand their role in health and disease, we must know very well their phenotype. Previous studies have identified and characterized EC mainly by immunohistochemistry, but there are also some studies using flow cytometry (FCM) after exposing these cells to enzymatic digestion, either to isolate and/ or to detach them from the vessel wall. However, it is well known that enzymatic treatment can cause deleterious effects on cell surface receptors, then influencing the antibody-antigen reaction. We describe a simple and cheap mechanical method to isolate EC from human vessels, avoiding alterations in the expression of cell surface receptors caused by the use of enzymes, and we tested it using FCM. With this method we were able to obtain fresh EC that were identified by FCM as a well-defined cluster of CD45-CD146+brightCD31+bright cells. This approach can be used in the future to isolate EC for further immunophenotypic characterization and for ex-vivo functional studies, as well as to test the effect of different stimuli, including pharmacological drugs.

Availability of sufficient quantity and quality of sugarcane planting materials from conventional seed source is one of the major challenges in the Ethiopian sugar estates. To circumvent this challenge, tissue culture technology is found to be the best alternative for which in vitro propagation protocol is a key pre-request. Thus, the present study was aimed to optimize protocol for in vitro rooting and acclimatization of two elite sugarcane genotypes i.e., N52 and N53. Experiments were laid out in a completely randomized design with factorial treatment arrangements. Half strength MS liquid media supplemented with combination of Sucrose (0, 40, 50, 60 and 70 g/l) and NAA (0,3,5 and 7 mg/l) along with two sugarcane genotypes (N52, N53) were used for rooting while substrate containing sand, soil and farmyard manure in six different ratios (1:1:0, 1:1:1, 1:2:1, 2:1:1, 1:1:2 and 1:2:0) were used for acclimatization. With regard to in vitro rooting, ½ strength liquid MS medium + 50 g/l sucrose + 3 mg/l NAA induced the highest rooting (100%) with 23.5 ± 1.29 average root number per shoot and 4.95 cm ± 0.06 cm root length in genotype N52 while 5 mg/l NAA + 50 g/l sucrose induced the highest (100%) rooting response with an average of 21.76 ± 0.57 root number per shoot with 4.54 cm ± 0.06 cm root length in sugarcane genotype N53. In acclimatization, best survival rate (94% in N52 and 100% in N53) was achieved on substrate mixtures containing sand + soil in 1:1: ratios. Thus, it can be deduced that this protocol can be used successfully for in vitro rooting and acclimatization of these genotypes.

Background: Many surgical strategies for reconstruction of both bone and cartilage have ever been investigated to restore joint structure and function in the late stages of Osteoarthritis (OA). This study was designed to investigate the regeneration of articular cartilage and subchondral bone in the loading-bearing site using a three-dimensional (3D) construct of autologous adipose tissue-derived mesenchymal stem cells (AT-MSCs).

Methods: A 3D construct consisting of approximately 1,920 spheroids each containing 5.0 × 104 AT-MSCs was implanted into an osteochondral defect (with a diameter of 6.8 mm and a depth of 6 mm) in the right femoral medial condyle in five adult mini-pigs. The contralateral (left femoral) defect was the control. At three and six months post-operatively, the defects were evaluated using both CT and MR imaging. The radiolucent volume (RV, mm3) of the defects was calculated based on the multiplanar reconstruction of the CT images. MR images and gross and histologic pathology features were scored using a modified-MOCART system and the ICRS system, respectively, at six months post-operatively.

Results: The percentages of RVs at three and six months compared with those immediately after the surgeries were significantly decreased in the implanted defects compared with the control defects. The total scores of modified- MOCART system were also significantly increased in the implanted sites comparing to the controls. Although there were no statistical differences in the average of gross scores, the average histological scores were significantly higher in the implanted sites than in the control sites.

Conclusion: This is the first report suggesting that implantation of a scaffold-free three dimensional construct of only AT-MSCs into the osteochondral defect regenerates the original cartilage and subchondral bone structures over six months post-operatively in the loading-bearing site of large animal.