Frozen-thawed gelatin-induced osteogenic cell sheets of canine adipose-derived mesenchymal stromal cells improved fracture healing in canine model

We assessed the efficacy of frozen-thawed gelatin-induced osteogenic cell sheet (FT-GCS) compared to that of fresh gelatin-induced osteogenic cell sheet (F-GCS) with adipose-derived mesenchymal stromal cells (Ad-MSCs) used as the control. The bone differentiation capacity of GCS has already been studied. On that basis, the experiment was conducted to determine ease of use of GCS in the clinic. In vitro evaluation of F-GCS showed 3–4 layers with an abundant extracellular matrix (ECM) formation; however, cryopreservation resulted in a reduction of FT-GCS layers to 2–3 layers. Cellular viabilities of F-GCS and FT-GCS did not vary significantly. Moreover, there was no significant difference in mRNA expressions of Runx2, β-catenin, OPN, and BMP-7 between F-GCS and FT-GCS. In an in vivo experiment, both legs of six dogs with transverse radial fractures were randomly assigned to one of three groups: F-GCS, FT-GCS, or control. Fracture sites were wrapped with the respective cell sheets and fixed with 2.7 mm locking plates and six screws. At 8 weeks after the operations, bone samples were collected and subjected to micro computed tomography and histopathological examination. External volumes of callus as a portion of the total bone volume in control, F-GCS, and FT-GCS groups were 49.6%, 45.3%, and 41.9%, respectively. The histopathological assessment showed that both F-GCS and FT-GCS groups exhibited significantly (p < 0.05) well-organized, mature bone with peripheral cartilage at the fracture site compared to that of the control group. Based on our results, we infer that the cryopreservation process did not significantly affect the osteogenic ability of gelatin-induced cell sheets.

Different Bone Healing Effects of Undifferentiated and Osteogenic Differentiated Mesenchymal Stromal Cell Sheets in Canine Radial Fracture Model

Cell sheets technology is being available for fracture healing. This study was performed to clarify bone healing mechanism of undifferentiated (UCS) and osteogenic (OCS) differentiated mesenchymal stromal cell (MSC) sheets in the fracture model of dogs. UCS and OCS were harvested at 10 days of culture. Transverse fractures at the radius of six beagle dogs were assigned into three groups (n = 4 in each group) i.e. UCS, OCS and control. The fractures were fixed with a 2.7 mm locking plate and six screws. Cell sheets were wrapped around the fracture site. Bones were harvested 8 weeks after operation, then scanned by micro-computed tomography (micro-CT) and analyzed histopathologically. The micro-CT revealed different aspects of bone regeneration among the groups. The percentages of external callus volume out of total bone volume in control, UCS, and OCS groups were 42.1, 13.0 and 4.9% (p < 0.05) respectively. However, the percentages of limbs having connectivity of gaps were 25, 12.5 and 75% respectively. In histopathological assessments, OCS group showed well organized and mature woven bone with peripheral cartilage at the fracture site, whereas control group showed cartilage formation without bone maturation or ossification at the fracture site. Meanwhile, fracture site was only filled with fibrous connective tissue without endochondral ossification and bone formation in UCS group. It was suggested that the MSC sheets reduced the quantity of external callus, and OCS induced the primary bone healing.

Elemental analysis of ten plant species at three phenological stages

Livestock rearing is a common practice in Tehsil Takht-e-Nasrati by the locals to earn their livelihood. Low productivity due to poor health of livestock is major consideration in the study area. In order to know whether this low productivity and poor health might be due to poor quality of forage, ten plant species were analyzed for micro mineral quantification at three phenological stages. Mineral compositions of plants are used to diagnose probable deficiencies in food and forage and nutritional status of plants. Comparison among at vegetative stage of test species and micro nutrients showed that the amount of manganese was found high [64.4mg Kg[-1]] in Chrozophora obliqua. Comparison among at reproductive and post reproductive stages of test species and micro nutrients pointed out that the amount of nickel was found high [67.24mg Kg[-1]] and [80.4mg Kg[-1]] respectively in Vitex trifolia. It is concluded that micro-minerals concentrations available in these forage plants to the grazing livestock were very low, hence this may be, one of the causes responsible for the pitiable health and productivity of the grazing animals in investigated area

Effect of phenological stages on nutritional assessment of ten plant species in Tehsil Takht-e-Nasrati, District Karak, Pakistan

The proximate analysis of ten plant species from Tehsil takht-e-Nasrati, district Karak was conducted at three phenological stages using standard techniques. The species showed variable results. In the test species, the moisture content varied from 2.11-14.12%, 1.85-11.51 and 1.12-11.51%; the ash contents varied from 3.08-17.16%, 5.09-18.12% and 6.13-19.09%; the fibre contents varied from 2.14-11.09%, 2.43-12.31% and 2.78-13.01%; the carbohydrate contents varied from 54-85.1%, [50-84.1] and 47.1-81.1%; the fat contents varied from 1.06-5.46%, 1.09-5.98% and 1.56- 6.57%; the protein contents varied from 2.11-12.41%, 3.14-15.87% and 4.12-18.54%; the energy value varied from 305.12- 394.23 KCal/100g, 314.21-423.13 KCal /100g and 321.23-434.26 KCal /100g at vegetative stage, reproductive stage and post reproductive stage respectively. Proximate analysis of such plant species can help us to determine the health benefits achieved from their use in society. It is concluded that the poor livestock productivity in research area is partially due to insufficient amount of available poor quality forage that requires attention of the range mangers to improve the habitat and livestock breeds