Wound healing potential of human umbilical cord mesenchymal stem cell conditioned medium: An in vitro and in vivo study in diabetes-induced rats.

BACKGROUND AND AIM: Human umbilical cord mesenchymal stem cells (hUC-MSCs) and its conditioned medium (CM) promote wound healing. This study investigated the wound healing potential of hUC-MSC CM in vitro and in vivo using diabetic animal models. MATERIALS AND METHODS: The CM from hUC-MSC CM prepared under hypoxic conditions (hypoxic hUC-MSC) was evaluated for stimulating rat fibroblast growth, collagen production (in vitro), and wound healing in animal models (in vivo). An excision wound on the dorsal side of the diabetes-induced rats was established, and the rats were randomly divided into non-treatment, antibiotic, and hypoxic hUC-MSC CM groups. The cell number of fibroblasts and collagen secretion was evaluated and compared among the groups in an in vitro study. By contrast, wound size reduction, width of re-epithelialization, and the collagen formation area were assessed and compared among the groups in an in vivo study. RESULTS: CM under hypoxic conditions contained a higher concentration of wound healing-related growth factors. Hypoxic hUC-MSC CM could facilitate fibroblast cell growth and collagen synthesis, although not significant compared with the control group. Re-epithelialization and collagen production were higher in the hUC-MSC CM group than in the antibiotic and non-treatment groups. CONCLUSION: Hypoxic hUC-MSC CM possessed more positive effects on the wound healing process based on re-epithelialization and collagen formation than antibiotic treatment did.

Motoric Recovery After Transplantation of Bone Marrow Derived Mesenchymal Stem Cells in Chronic Spinal Cord Injury: A Case Report.

BACKGROUND For the past 20 years, numerous of clinical trials focusing on the use of mesenchymal stem cells (MSC) in spinal cord injury (SCI) treatment has been conducted. However, controversies over whether stem cells are the main factor in a patient's recovery still persisted in sub-acute SCI. This study aimed to evaluate the motoric recovery in a chronic SCI patient treated with bone marrow derived MSC (BM-MSC) transplantation. CASE REPORT We present a case report of patient with a 12-year-long-chronic SCI that was treated by BM-MSC) transplantation using a serial administration protocol. The protocol consisted of direct parenchymal injection to the affected lesion and multiple (5 times) intravenous stem cell injection as the adjuncts. There was no complication or serious adverse effects encountered during the procedure and follow up. At the final follow up of 5 years, the patient neurological status improved from American Spinal Injury Association (ASIA) A status to ASIA C status, which signifies improvement in his ambulatory status. Magnetic resonance imaging and electrophysiology examination also showed changes that indicated recovery of the neurologic function. CONCLUSIONS Based on the limited adverse reaction and outcome, our case report may serve as an additional alternative protocol in stem cell administration to improve the outcome of chronic spinal cord injury patients.

Autologous mesenchymal stem cell implantation, hydroxyapatite, bone morphogenetic protein-2, and internal fixation for treating critical-sized defects: a translational study.

INTRODUCTION: Critical-sized defect (CSD) is one of the most challenging cases for orthopaedic surgeons. We aim to explore the therapeutic potential of the combination of bone marrow-derived mesenchymal stem cells (BM-MSCs), hydroxyapatite (HA) granules, bone morphogenetic protein-2 (BMP-2), and internal fixation for treating CSDs. METHODS: This was a translational study performed during the period of January 2012 to 2016. Subjects were patients diagnosed with CSDs who had previously failed surgical attempts. They were treated with the combination of autologous BM-MSCs, HA granules, BMP-2, and mechanical stabilization. Post-operative pain level, functional outcome, defect volume, and radiological healing were evaluated after a minimum follow-up of 12 months. RESULTS: A total of six subjects were recruited in this study. The pain was significantly reduced in all cases; with the decrease of mean preoperative visual analog scale (VAS) from 4 ± 2.2 to 0 after six month follow-up. Clinical functional outcome percentage increased significantly from 25 ± 13.7 to 70.79 ± 19.5. Radiological healing assessment using Tiedemann score also showed an increase from 0.16 ± 0.4 to 8 ± 3 at one year follow-up. No immunologic nor neoplastic side effects were found. CONCLUSIONS: The combination of autologous BM-MSCs, HA granules, and BMP-2 is safe and remains to be a good option for the definitive treatment for CSD with previous failed surgical attempts. Further studies with a larger sample size are required to be done.

Progenitor Hematopoietic Cells Implantation Improves Functional Capacity of End Stage Coronary Artery Disease Patients with Advanced Heart Failure.

Background. Proangiogenic Hematopoietic Cells (PHC) which comprise diverse mixture of cell types are able to secrete proangiogenic factors and interesting candidate for cell therapy. The aim of this study was to seek for benefit in implantation of PHC on functional improvement in end stage coronary artery disease patients with advanced heart failure. Methods. Patients with symptomatic heart failure despite guideline directed medical therapy and LVEF less than 35% were included. Peripheral blood mononuclear cells were isolated, cultivated for 5 days, and then harvested. Flow cytometry and cell surface markers were used to characterize PHC. The PHC were delivered retrogradely via sinus coronarius. Echocardiography, myocardial perfusion, and clinical and functional data were analyzed up to 1-year observation. Results. Of 30 patients (56.4 ± 7.40 yo) preimplant NT proBNP level is 5124.5 ± 4682.50 pmol/L. Harvested cells characterized with CD133, CD34, CD45, and KDR showed 0.87 ± 0.41, 0.63 ± 0.66, 99.00 ± 2.60, and 3.22 ± 3.79%, respectively. LVEF was improved (22 ± 5.68 versus 26.8 ± 7.93, p < 0.001) during short and long term observation. Myocardial perfusion significantly improved 6 months after treatment. NYHA Class and six-minute walk test are improved during short term and long term follow-up. Conclusion. Expanded peripheral blood PHC implantation using retrograde delivery approach improved LV systolic function, myocardial perfusion, and functional capacity.

Effect of Staphylococcus aureus and Staphylococcus epidermidis debris and supernatant on bone marrow stromal cells growth.

AIM: the goal of this study is to observe the effect of SA and SE debris and supernatant on BMSCs growth. METHODS: SA and SE were isolated and cultured from lesion materials of spondylitis patients which were determined by gram staining and biochemical tests. BMSCs were cultured and incubated for 11 days to be further sub-cultured and tripsinized for cell counting before seeded. 0.1 mg/ml SA and SE debris and supernatant were added into the BMSCs culture media. Cell counting was performed 2, 5, 7, and 9 days after debris and supernatant addition to get the growth profile of BMSCs. RESULTS: debris and supernatant decreases BMSCS number at initial day. At day 5, BMSCs in the group debris were growing down, mean while BMSCs in the group supernatant were able to retain the cell number. Viability of all groups was more than 80%. CONCLUSION: both debris and supernatant from SA and SE have inhibitory effect of the growth BMSCs in the initial day. BMSCs could provide barrier to survive from the debris and supernatant environment of both bacteria in day five. BMSCs can use supernatant to retain the growth to replace the lack of nutrition.

Isolation and cultivation of mesenchymal stem cells from iliac crest bone marrow for further cartilage defect management.

AIM: to validate isolation and cultivation methods of bone marrow mesenchymal stem cells (BM-MSCs) from iliac crest, and to compare biological characteristics of BM-MSCs from different age groups for preparation of autologous stem cell therapy in cartilage defect. METHODS: patients undergoing spinal surgery were selected and grouped according to age. Iliac crest bone marrow from the patients was aspirated. BM-MSCs were isolated from the bone marrow and then cultivated. Their biological characteristics including morphological appearances and surface biomarkers were evaluated. Growth curves were observed. Sterility and Mycoplasma tests were also performed for quality assessment of BM-MSCs culture procedure. RESULTS: in average, cultivated-BM-MSCs reached the number of 7.56-22.95 x 106 in 4-7 weeks period. BM-MSCs of all age groups showed the same quality of morphology, shape and surface biomarkers (CD105+, CD73+, CD34-, CD45-, CD14-, CD19-, HLA-DR-). CONCLUSION: our procedures in isolating and cultivating of BM-MSCs have reached required amount for implantation into the cartilage lesion. In addition, the cultivated-BM-MSCs' biological characteristics were also in accordance with International Society of Cell Therapy (ISCT) MSCs criteria.

Enzymatic synthesis of two novel non-reducing oligosaccharides using transfructosylation activity with beta-fructofuranosidase from arthrobacter globiformis.

Two novel non-reducing oligosaccharides together with tri- and tetra-saccharides were synthesized by transfructosylation activity from sucrose as a donor and cellobiose or cellotriose as an acceptor with a purified beta-fructofuranosidase from Arthrobacter globiformis IFO 3062, and these oligosaccharides were identified as O-beta-D-glucopyranosyl-(1-->4)-alpha-D-glucopyranosyl-(1-->2)-alpha,beta-D-fructofuranoside and O-beta-D-glucopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->4)-alpha-D-glucopyranosyl-(1-->2)-alpha,beta-D-fructofuranoside by spectrometric analyses. Both oligosaccharides were stable under condition at 100 degrees C for 30 min, and showed no degradation at pH 2.

Cloning and heterologous expression of a beta-fructofuranosidase gene from Arthrobacter globiformis IFO 3062, and site-directed mutagenesis of the essential aspartic acid and glutamic acid of the active site.

We have cloned the gene encoding a beta-fructofuranosidase from Arthrobacter globiformis IFO 3062, and subsequently, the gene was heterologously expressed in Escherichia coli. This beta-fructofuranosidase gene encodes a protein of 548 amino acid residues with a calculated molecular mass of 60,519 Da. We have examined the roles of three residues of A. globiformis IFO 3062 beta-fructofuranosidase by site-directed mutagenesis, and found that aspartic acid 130 and glutamic acid 392, which are two of the apparent catalytic residues, are essential for hydrolase activity. This study provides the first experimental evidence showing that these two amino acid residues of beta-fructofuranosidase play a critical role in hydrolyzing sucrose.