Partial Purification and Characterization of Two Endo-ȕ-1, 4-glucanase from Trichoderma sp. (Shmosa tri)

Abstract: Two endoglucanases (EC 3.2.1.4) from Trichoderma sp. (shmosaTri) FJ937359 were purified to homogeneity using ammonium sulfate precipitation and gel filtration. Purity was confirmed by SDS/PAGE. Enzymatic properties and molecular weights were determined. Molecular weights of CMCase I and II were 58 and 34 KDa, respectively. The effect of temperature on the 2 endoglucanase activity was studied and results showed that optimum activity obtained at 50°C for both CMCase I and II. The enzymes withstand 60 min at 50ºC without loss of enzymatic activity. CMCase I and II retained 14.0 and 26.5 % of their original activities at 70°C after 90 min. The optimum pH for CMCase I and II was 5.0. Results also show that CMCase I was active at room temperature after 24 hrs over a broad pH range (3.0-9.0) while CMCase II was relatively stable in pH range (4.0-6.0). Among different kinds of substrates, both enzymes showed a high preference for carboxymethyl cellulose while both CMCase I and II did not show any hydrolytic activity against chitin, starch and cellobiose. On the other hand both CMCase I and II have relatively low hydrolytic activity towards ȕ glucan and xylan. All metallic ions used as well as EDTA and SDS at a concentration of 20 ug/ml of reaction mixture have an inhibitory effect on both CMCase I and II

The Biomechanical Effect of Different Denture Base Materials on the Articular Disc in Complete Denture Wearers: A Finite Element Analysis

AIM: The objective of the present study was to evaluate the effect of different denture base materials on the stress distribution in TMJ articular disc (AD) in complete denture wearers.MATERIAL AND METHODS: Two three dimensional Finite Element (FEA) models of an individual temporomandibular joint (TMJ) were built on the basis CT scan. The FEA model consisted of four parts: the condyle, the articular disc, the denture base, and the articular eminence skull. Acrylic resin and chrome-cobalt denture base materials were studied. Static loading of 300N was vertically applied to the central fossa of the mandibular second premolar. Stress and strain were calculated to characterize the stress/strain patterns in the disc.RESULTS: The maximum tensile stresses were observed in the anterior and posterior bands of (AD) on load application with the two denture base materials. The superior boundaries of the glenoid fossa showed lower stress than those on the inferior boundaries facing the condyle.CONCLUSIONS: Within the limitations of the present study it may be concluded that: The denture base material may a have an effect in stress-strain pattern in TMJ articular disc. The stiffer denture base material, the better the distribution of the load to the underling mandibular supporting structures & reducing stresses induced in the articular disc

Combination of Human Amniotic Fluid Derived-Mesenchymal Stem Cells and Nano-hydroxyapatite Scaffold Enhances Bone Regeneration

BACKGROUND: Human amniotic fluid-derived stem cells (hAF-MSCs) have a high proliferative capacity and osteogenic differentiation potential in vitro. The combination of hAF-MSCs with three-dimensional (3D) scaffold has a promising therapeutic potential in bone tissue engineering and regenerative medicine. Selection of an appropriate scaffold material has a crucial role in a cell supporting and osteoinductivity to induce new bone formation in vivo. AIM: This study aimed to investigate and evaluate the osteogenic potential of the 2nd-trimester hAF-MSCs in combination with the 3D scaffold, 30% Nano-hydroxyapatite chitosan, as a therapeutic application for bone healing in the induced tibia defect in the rabbit. SUBJECT AND METHODS: hAF-MSCs proliferation and culture expansion was done in vitro, and osteogenic differentiation characterisation was performed by Alizarin Red staining after 14 & 28 days. Expression of the surface markers of hAF-MSCs was assessed using Flow Cytometer with the following fluorescein-labelled antibodies: CD34-PE, CD73-APC, CD90-FITC, and HLA-DR-FITC. Ten rabbits were used as an animal model with an induced defect in the tibia to evaluate the therapeutic potential of osteogenic differentiation of hAF-MSCs seeded on 3D scaffold, 30% Nano-hydroxyapatite chitosan. The osteogenic differentiated hAF-MSCs/scaffold composite system applied and fitted in the defect region and non-seeded scaffold was used as control. The histopathological investigation was performed at 2, 3, & 4 weak post-transplantation and scanning electron microscope (SEM) was assessed at 2 & 4 weeks post-transplantation to evaluate the bone healing potential in the rabbit tibia defect. RESULTS: Culture and expansion of 2nd-trimester hAF-MSCs presented high proliferative and osteogenic potential in vitro. Histopathological examination for the transplanted hAF-MSCs seeded on the 3D scaffold, 30% Nano-hydroxyapatite chitosan, demonstrated new bone formation in the defect site at 2 & 3 weeks post-transplantation as compared to the control (non-seeded scaffold). Interestingly, the scaffold accelerated the osteogenic differentiation of AF-MSCs and showed complete bone healing of the defect site as compared to the control (non-seeded scaffold) at 4 weeks post-transplantation. Furthermore, the SEM analysis confirmed these findings. CONCLUSION: The combination of the 2nd-trimester hAF-MSCs and 3D scaffold, 30% Nano-hydroxyapatite chitosan, have a therapeutic perspective for large bone defect and could be used effectively in bone tissue engineering and regenerative medicine

Osteogenic Differentiation Potential of Human Bone Marrow and Amniotic Fluid-Derived Mesenchymal Stem Cells in Vitro & in Vivo

BACKGROUND: Cell therapies offer a promising potential in promoting bone regeneration. Stem cell therapy presents attractive care modality in treating degenerative conditions or tissue injuries. The rationale behind this is both the expansion potential of stem cells into a large cell population size and its differentiation abilities into a wide variety of tissue types, when given the proper stimuli. A progenitor stem cell is a promising source of cell therapy in regenerative medicine and bone tissue engineering. AIM: This study aimed to compare the osteogenic differentiation and regenerative potentials of human mesenchymal stem cells derived from human bone marrow (hBM-MSCs) or amniotic fluid (hAF-MSCs), both in vitro and in vivo studies. SUBJECTS AND METHODS: Human MSCs, used in this study, were successfully isolated from two human sources; the bone marrow (BM) and amniotic fluid (AF) collected at the gestational ages of second or third trimesters. RESULTS: The stem cells derived from amniotic fluid seemed to be the most promising type of progenitor cells for clinical applications. In a pre-clinical experiment, attempting to explore the therapeutic application of MSCs in bone regeneration, Rat lumbar spines defects were surgically created and treated with undifferentiated and osteogenically differentiated MSCs, derived from BM and second trimester AF. Cells were loaded on gel-foam scaffolds, inserted and fixed in the area of the surgical defect. X-Ray radiography follows up, and histopathological analysis was done three-four months post- operation. The transplantation of AF-MSCs or BM-MSCs into induced bony defects showed promising results. The AF-MSCs are offering a better healing effect increasing the likelihood of achieving successful spinal fusion. Some bone changes were observed in rats transplanted with osteoblasts differentiated cells but not in rats transplanted with undifferentiated MSCs. Longer observational periods are required to evaluate a true bone formation. The findings of this study suggested that the different sources; hBM-MSCs or hAF-MSCs exhibited remarkably different signature regarding the cell morphology, proliferation capacity and osteogenic differentiation potential CONCLUSIONS: AF-MSCs have a better performance in vivo bone healing than that of BM-MSCs. Hence, AF derived MSCs is highly recommended as an alternative source to BM-MSCs in bone regeneration and spine fusion surgeries. Moreover, the usage of gel-foam as a scaffold proved as an efficient cell carrier that showed bio-compatibility with cells, bio-degradability and osteoinductivity in vivo

Significance of Residual Organisms in Foods after Substerilizing Doses of Gamma Radiation: A Review

The nature of the flora surviving substerilizing irradiation of foods is dependent on the dose applied and the conditions of applications as well as the microenvi¬ronment of the food. At a dose level low enough to preserve acceptable sensory properties of fresh food, few if any of the common contaminants of public health significance survive irradiation. Survivors are weakened and present no unique problem of acquired resistance through recycling. Those bacteria surviving a low dose treatment of a relatively contaminated product like ground beef can grow under normal storage conditions and cause obvious sensory spoilage. Thus, low dose irradiation extends the shelf-life of fresh foods and reduces public health hazards, but foods so treated require normal care in production and distribution

Pyrvinium Targets the Unfolded Protein Response to Hypoglycemia and Its Anti-Tumor Activity Is Enhanced by Combination Therapy

We identified pyrvinium pamoate, an old anthelminthic medicine, which preferentially inhibits anchorage-independent growth of cancer cells over anchorage-dependent growth (∼10 fold). It was also reported by others to have anti-tumor activity in vivo and selective toxicity against cancer cells under glucose starvation in vitro, but with unknown mechanism. Here, we provide evidence that pyrvinium suppresses the transcriptional activation of GRP78 and GRP94 induced by glucose deprivation or 2-deoxyglucose (2DG, a glycolysis inhibitor), but not by tunicamycin or A23187. Other UPR pathways induced by glucose starvation, e.g. XBP-1, ATF4, were also found suppressed by pyrvinium. Constitutive expression of GRP78 via transgene partially protected cells from pyrvinium induced cell death under glucose starvation, suggesting that suppression of the UPR is involved in pyrvinium mediated cytotoxicity under glucose starvation. Xenograft experiments showed rather marginal overall anti-tumor activity for pyrvinium as a monotherapy. However, the combination of pyrvinium and Doxorubicin demonstrated significantly enhanced efficacy in vivo, supporting a mechanistic treatment concept based on tumor hypoglycemia and UPR