The future of bone marrow stromal cell transplantation for the treatment of spinal cord injury

Bone marrow stromal cell （BMSC） transplantation therapy is a promising approach for treating spinal cord injury （SCI）, based on a number of experimental and clinical reports （Wright et al., 2011）. BMSCs are a source of neuroregenerative somatic stem cells that are without the potential for tumorigenicity. Although clinical studies of autologous BMSC transplantation have been reported in Asia （fiang et al., 2013; Yoon et al., 2007）, in Japan, it is currently an uncommon procedure and highly controversial as well. This perspective paper provides an overview of the clinical effectiveness of BMSC trans- 191antation and a proposal to enhance its use as a viable therapy.

S100 protein expression during induced Schwann cell-like cell differentiation of rat bone marrow mesenchymal cells in vitro

BACKGROUND： S100 protein can promote axonal growth. Therefore, transplantation of induced bone marrow-derived mesenchymal stem cells （MSCs） that can secrete S100 may provide a beneficial microenvironment for neural regeneration. OBJECTIVE： To explore the changes in S100 expression during rat MSCs differentiation into Schwann ceils in vitro. DESIGN, TIME AND SETTING： This cytology experiment was performed at the Jiangsu Key Laboratory of Neuroregeneration, Nantong University in China, from January 2006 to May 2007. MATERIALS： The rabbit anti-S100 polyclonal antibody was purchased from Dako, Denmark; the mouse anti-rat S100 monoclonal antibody was purchased from Sigma, USA. METHODS： MSCs were cultured from adult Sprague-Dawley rat femur and tibia. Cell proliferation was determined by the MTT method and CD markers, and cell cycle was measured by flow cytometry. MSCs were induced to differentiate into SC cells. SC cells were stained for S100 protein, glial fibrillary acidic protein, and low-affinity nerve growth factor receptor. S100 protein and mRNA levels were evaluated by flow cytometry, Western blot, and reverse transcription-polymerase chain reaction. MAIN OUTCOME MEASURES： S100 protein and mRNA expression. RESULTS： MSCs exhibited high amplification potential over eight passages. Prior to induction, the majority of MSCs were at the G0/G1 phase of the cell cycle. After induction, MSCs displayed morphology changes similar to Schwann cells. Moreover, induction increased S100 mRNA levels. Immunofluorescence showed that MSCs expressed S100 protein, glial fibrillary acidic protein, and low-affinity nerve growth factor receptor at 7 days of induction. Induction also increased S100 protein levels compared with untreated MSCs. CONCLUSION： MSCs are capable of differentiating into Schwann cells-like cells under conditional induction in vitro, with increasing S100 mRNA and protein expression.

Synergistic effects of brain-derived neurotrophic factor and retinoic acid on inducing the differentiation of bone marrow stromal cells into neuron-like cells in adult rats in vitro

BACKGROUND： Under induction of retinoic acid （RA）, bone marrow stromal cells （BMSCs） can differentiate into nerve cells or neuron-like cells, which do not survive for a long time, so those are restricted to an application. Other neurotrophic factors can also differentiate into neuronal cells through inducing BMSCs; especially, brain-derived neurotrophic factor （BDNF） can delay natural death of neurons and play a key role in survival and growth of neurons. The combination of them is beneficial for differentiation of BMSCs. OBJECTIVE： To investigate the effects of BDNF combining with RA on inducing differentiation of BMSCs to nerve cells of adult rats and compare the results between common medium group and single BDNF group. DESIGN： Randomized controlled animal study SETTING： Department of Neurology, Affiliated Hospital of Xuzhou Medical College MATERIALS： The experiment was carried out in the Clinical Neurological Laboratory of Xuzhou Medical College from September 2003 to April 2005. A total of 24 SD rats, of either gender, 2 months old, weighing 130-150 g, were provided by Experimental Animal Center of Xuzhou Medical College [certification： SYXK （su） 2002-0038]. Materials and reagents： low-glucose DMEM medium, bovine serum, BDNF, RA, trypsin, separating medium of lymphocyte, monoclonal antibody of mouse-anti-nestin, neuro-specific enolase, glial fibrillary acidic protein （GFAP） antibody, SABC kit, and diaminobenzidine （DAB） color agent. All these mentioned above were mainly provided by SIGMA Company, GIBCO Company and Boshide Company. METHODS： Bone marrow of SD rats was selected for density gradient centrifugation. BMSCs were undertaken primary culture and subculture; and then, those cells were induced respectively in various mediums in total of 3 groups, including control group （primary culture）, BDNF group （20 μg/L BDNF） and BDNF＋RA group （20 μg/L BDNF plus 20 μg/L RA）. On the 3^rd and the 7^th days after induction, BMSCs were stained immunocytochemically with nestin （sign of nerve stem cells）, neuron-specific enolase （NSE, sign of diagnosing neurons） and GFAP （diagnosing astrocyte）, and evaluated cellular property. MAIN OUTCOME MEASURES ： Induction and differentiation in vitro of BMSCs in 3 groups RESULTS： （1） Induction and differentiation of BMSCs： Seven days after induction, cells having 2 or more apophyses were observed. Soma shaped like angle or erose form, which were similar to neurons and glial cells having strong refraction. （2） Results of immunocytochemical detection： Three days after induction, rate of positive cells in BDNF＋RA group was higher than that in BDNF group and control group [（86.15±4.58）%, （65.43±4.23）%, （4.18±1.09）%, P 〈 0.01]. Seven days after induction, rate of positive cells was lower in BDNF group and BDNF＋RA group than that in both groups at 3 days after induction [（31.12±3.18）%, （29.35±2.69）%, P 〈 0.01]; however, amounts of positive cells of NSE and GFAP were higher than those at 3 days after induction （P 〈 0.01）; meanwhile, the amount in BDNF＋RA group was remarkably higher than that in BDNF group （P 〈 0.01）. CONCLUSION： Combination of BDNF and RA can cooperate differentiation of BMSCs into neurons and astrocyte, and the effect is superior to single usage of BDNF.

PURGE OF MALIGNANT CELLS FROM BONE MARROW BY HEMATOPORPHYRIN DERIVATIVES AND LIGHT EXPOSURE IN VITRO

During autologous bone marrow graft in treatment of malignant diseases, it is critical to purge malignant cells from the marrow. In the present study, the sensitivity to photodynamic inactivation of 3 leukemic cell lines was compared with their counterpart normal hematopoietic cells. After mouse leukemic L1210 cells were treated with a preparation of hematoporphyrin derivatives, YHpD, 10 μg/ml for 1 hr. and irradiated with blacklight (peak wavelength 395 nm, light intensity 0.6 mW/cm2) for 5 minutes, the survival rate of clonogenic cells decreased to <10%, while that of bone marrow granulocyte macrophage progenitor cells (CFU-GM) in DBA/2 mice remained at nearly normal level (>80%). Similar results were obtained when human leukemic HL-60 cells were compared with human CFU-GM and mouse leukemic L615 cells with CFU-GM in 615 strain mice. It is suggested that hematoporphyrin photoradiation may be useful for Iselectively killing leukemic cells in bone marrow.

Effect of L-lysine in culture medium on nodule formation by bone marrow cells

The aim of this study was to estimate the effect of L-lysine on nodule formation by rat bone marrow cells in vitro. In this study, L-lysine was added to medium for mesenchymal stem cell culture to promote proliferation and differentiation of the cells, and then nodule formation was estimated in an in vitro rat bone marrow cell culture. Bone marrow cells from the bone shafts of the femora of Fischer 344 rats were cultured in minimum essential medium with 20 μl of L-lysine solution at 10﹣4, 10﹣5, 10﹣6, 10﹣7 or 10﹣8 M. Dexamethasone was also added to the medium at 10 nM for differentiation of stem cells from bone marrow into osteoblast progenitor cells. The subculture was performed for 2 weeks. The quantity of osteocalcin in rat bone marrow cell culture with dexamethasone was 392 ng/ml. In the medium with dexamethasone and 10﹣8 M L-lysine, the quantity of osteocalcin was 437 ng/ml. Nodules only formed upon addition of 20 μl of L-lysine at 10﹣8 M. It was indicated that 10﹣8 M L-lysine should be the optimal concentration for calcification. For nodule formation by rat bone marrow cells in vitro, the optimum concentration of L-lysine in culture medium might be 20 μl of 10﹣8 M. L-lysine could play an important role in matrix production for bone formation in vitro.

Treatment of Unicameral and Aneurysmal Bone Cysts by Minimally Invasive Percutaneous Injection of Grafton DBF Putty Using the Kyphon Cement Delivery System

Background: Simple Unicameral and Aneurysmal Bone Cysts are benign lesions that may heal spontaneously especially after fracture which may be the first symptom. However, often size increases causing pain, and complications of fractures can severely compromise the patient. Aim: The results in a series of cases treated minimally invasive using a new device for the application of allogenic bone material appear highly promising and shall be presented. Patients and Methods: Eight consecutive patients with symptomatic Unicameral Bone Cysts (UBC) were treated by percutaneous instillation of Grafton? DBF Putty (demineralised allogenic bone containing fibers) mixed with autologous bone marrow using the Kyphon? Cement Delivery System (Medtronic), which allows the injection of this high viscosity paste by controlled high pressure. Five patients with Aneurysmal Bone Cysts (ABC) were treated accordingly after inactivation by Aethoxysclerol 3% and lacking bone formation. Using this approach a high rate of bone regeneration was observed in these patients at 8 months to 5 years follow-up (f/u). Conclusion: The presented technique of a minimally invasive biologic treatment led to highly satisfying results using the Grafton? DBF Putty with its higher potential for bone regeneration than demineralized bone matrix not containing fibres (DBM).

Stem Cell Ophthalmology Treatment Study(SCOTS) for retinal and optic nerve diseases: a case report of improvement in relapsing auto-immune optic neuropathy

We present the results from a patient with relapsing optic neuropathy treated within the Stem Cell Ophthalmology Treatment Study （SCOTS）. SCOTS is an Institutional Review Board ap- proved clinical trial and has become the largest ophthalmology stem cell study registered at the National Institutes of Health to date （www.clinicaltrials.gov Identifier NCT 01920867）. SCOTS utilizes autologous bone marrow-derived stem cells （BMSCs） for treatment of retinal and optic nerve diseases. Pre-treatment and post-treatment comprehensive eye exams of a 54 year old female patient were performed both at the Florida Study Center, USA and at The Eye Center of Columbus, USA. As a consequence of a relapsing optic neuritis, the patient＇s previously normal visual acuity decreased to between 20/350 and 20/400 in the right eye and to 20/70 in the left eye. Significant visual field loss developed bilaterally. The patient underwent a right eye vitrectomy with injection of BMSCs into the optic nerve of the right eyeand retrobulbar, subtenon and in- travitreal injection of BMSCs in the left eye. At 15 months after SCOTS treatment, the patient＇s visual acuity had improved to 20/150 in the right eye and 20/20 in the left eye. Bilateral visual fields improved markedly. Both macular thickness and fast retinal nerve fiber layer thickness were maximally improved at 3 and 6 months after SCOTS treatment. The patient also reduced her mycophenylate dose from 1,500 mg per day to 500 mg per day and required no steroid pulse therapy during the 15-month follow up.

Pre-degenerated peripheral nerves co-cultured with bone marrow-derived cells: a new technique for harvesting high-purity Schwann cells

Schwann cells play an important role in the peripheral nervous system, especially in nerve repair following injury, so artificial nerve regen- eration requires an effective technique for obtaining purified Schwann cells. In vivo and in vitro pre-degeneration of peripheral nerves have been shown to obtain high-purity Schwann cells. We believed that in vitro pre-degeneration was simple and controllable, and available for the clinic. Thus, we co-cultured the crushed sciatic nerves with bone marrow-derived cells in vitro. Results demonstrated that, 3 hours after injury, a large number of mononuclear cells moved to the crushed nerves and a large number of bone marrow-derived cells infiltrated the nerve segments. These changes promoted the degradation of the nerve segments, and the dedifferentiation and proliferation of Schwann cells. Neural cell adhesion molecule and glial fibrillary acidic protein expression were detected in the crushed nerves. Schwann cell yield was 9.08 ± 2.01 ×104/mg. The purity of primary cultured Schwann cells was 88.4 ± 5.79%. These indicate a successful new method for ob- taining Schwann cells of high purity and yield from adult crushed sciatic nerve using bone marrow-derived cells.

Modified technique of advanced core decompression for treatment of femoral head osteonecrosis

BACKGROUND Osteonecrosis of the femoral head(ONFH)is a destructive condition most commonly affecting young and middle-aged patients.The leading consequence of ONFH is often a significant articular disability.Effective joint-preserving surgical treatments are urgently needed for patients with early stage ONFH when outcomes of treatment are in general better than the advanced stage disease.AIM To introduce a new surgery procedure called percutaneous expanded core decompression and mixed bone graft technique,which is a new way of jointpreserving surgical treatments.METHODS The clinical data of 6 patients with ONFH diagnosed and treated with the procedure called percutaneous expanded core decompression and mixed bone graft technique at The First Hospital of Qiqihar from March 2013 to August 2019 were retrospectively analyzed;the follow-up ended in December 2019.RESULTS There were 6 male patients with an average age of 43 years in our study.Gratifying results have been obtained from the comparison of Harris hip score,visual analogue scale,and imaging examination before and after operation.CONCLUSION This new modified technique is simple,safe,and reliable.No serious perioperative complications were observed in our cases.Advantages of the single blade expandable reamer are obvious.The adjuvant substance is inexpensive and easy to obtain.Thus,this technique is an effective joint-preserving surgical treatment for patients with early stage of ONFH.

Calcified Nodule Formation <i>in Vitro</i>Using Rat Mandibular Incisor Pulp Cells

One great advantage of bone marrow is that a large number of stem cells can be obtained. However, stem cells cannot be taken from the bone marrow of a patient for the purpose of regenerating teeth. In dental practice, it may not be practical to obtain mesenchymal stem cells for the regeneration of the tooth from iliac bone marrow, as in orthopedics. Therefore, establishment of a cell source for tooth regeneration is one of the important problems in the field of regenerative medicine. Although the isolation of undifferentiated mesenchymal stem cells or odontoblasts from dental pulp may be difficult, dental pulp may be a favorable source for stem cells to regenerate a tooth via tissue engineering. As a fundamental study for tooth regeneration, this study was performed in order to clarify the culture periods for proliferation and differentiation using rat dental pulp-derived cells. The results of this study were as follows: Primary culture of the dental pulp cells does not require a long period of time. However, for sufficient proliferation of dental pulp cells to form a calcified nodule in the secondary culture, a long culture period is required. Dexamethasone was essential for the formation of calcified nodules by dental pulp-derived cells. Calcified nodule formation by dental pulp-derived cells in vitro required more than one month. The duration of the secondary culture for the dental pulp-derived cells will be much longer.

Development of bone marrow mesenchymal stem cell culture in vitro

Objective To review the in vitro development of bone marrow mesenchymal stem cells culture （BM-MSC）.Data sources The data cited in this review were mainly obtained from articles listed in Medline and PubMed. The search terms were “bone marrow mesenchymal stem cell” and “cell culture”.Study selection Articles regarding the in vitro development of BM-MSCs culture, as well as the challenge of optimizing cell culture environment in two-dimensional （2D） vs. 3D.Results Improving the culture conditions increases the proliferation and reduces the differentiation. Optimal values for many culture parameters remain to be identified. Expansion of BM-MSCs under defined conditions remains challenging, including the development of optimal culture conditions for BMSC and large-volume production systems.Conclusions Expansion of BM-MSCs under defined conditions remains challenges, including the development of optimal culture conditions for BMSC and scale-up to large-volume production systems. Optimal values for many culture parameters remain to be identified.

Treatment of Bone Marrow Failure Syndrome with Integrated Traditional and Western Medicine

Aplastic anemia （AA） and myelodysplastic syndrome （MDS） are both included in the bone marrow failure syndromes （BMFS）. AA is a group of diseases characterized by hematopoietic stem/progenitor cell damage, oerioheral blood cvtooenia, andclinical manifestations including anemia, bleeding and infection, which eventually lead to bone marrow failure. The incidence rate of AA in China is 7.4/10^6, higher than that in Western countries, among which the morbidity of acute AA and chronic AA （CAA） is 1.4/10^6 and 6.0/10^6, respectively.

TREATMENT OF APLASTIC ANEMIA: ALLOGENEIC BONE MARROW TRANSPLANTATION VERSUS IMMUNOMODULATION THERAPY WITH ANTILYMPHOCYTE GLOBULIN

There are two arms in the management of aplastic anemia,one allogeneic bone marrow transplantation(BMT) and the other immunomodulation therapy with antilymphocyte globulin(ALG).

Clinical Study of the Treatment Based Mainly on Syndrome Differentiation for the Complications of Bone Marrow Transplantation in 22 Patients with Leukemia

Objective: To investigate the results of treatment of complications with traditional Chinese medicine (TCM) after bone marrow transplantation (BMT) in leukemic patients.Methods: Chinese herbal medicine was taken orally according to Syndrome Differentiation.Results: Ten patients with long-term fever (9 patients with unknown causes, 1 patient with lung infection) were cured, who were irresponsive to the combination of multi-antibiotics. Out of 3 patients with severe jaundice, it was cured in 1 patient with hepatic venoocclusive disease (VOD), and improved after treatment of TCM and deteriorated secondary to severe infection in the other 2 patients with graft versus-host-disease. Out of 9 patients with gastrointestinal side effects induced by high-dose radiotherapy or chemotherapy, it was cured in 6 patients, improved in 2 patients and no response in 1 patient.Conclusion: TCM was showed to be effective in treating the complications post-BMT in patients with leukemia, and could accelerate the recovery of patients.

In vitro cartilage production using an extracellular matrix-derived scaffold and bone marrow-derived mesenchymal stem cells

Background Cartilage repair is a challenging research area because of the limited healing capacity of adult articular cartilage.We had previously developed a natural,human cartilage extracellular matrix （ECM）-derived scaffold for in vivo cartilage tissue engineering in nude mice.However,before these scaffolds can be used in clinical applications in vivo,the in vitro effects should be further explored.Methods We produced cartilage in vitro using a natural cartilage ECM-derived scaffold.The scaffolds were fabricated by combining a decellularization procedure with a freeze-drying technique and were characterized by scanning electron microscopy （SEM）,micro-computed tomography （micro-CT）,histological staining,cytotoxicity assay,biochemical and biomechanical analysis.After being chondrogenically induced,the induction results of BMSCs were analyzed by histology and Immunohisto-chemistry.The attachment and viability assessment of the cells on scaffolds were analyzed using SEM and LIVE/DEAD staining.Cell-scaffold constructs cultured in vitro for 1 week and 3 weeks were analyzed using histological and immunohistochemical methods.Results SEM and micro-CT revealed a 3-D interconnected porous structure.The majority of the cartilage ECM was found in the scaffold following the removal of cellular debris,and stained positive for safranin O and collagen Ⅱ.Viability staining indicated no cytotoxic effects of the scaffold.Biochemical analysis showed that collagen content was （708.2±44.7）μg/mg,with GAG （254.7±25.9） μg/mg.Mechanical testing showed the compression moduli （E） were （1.226±0.288） and （0.052±0.007） MPa in dry and wet conditions,respectively.Isolated canine bone marrow-derived stem cells （BMSCs） were induced down a chondrogenic pathway,labeled with PKH26,and seeded onto the scaffold.Immunofluorescent staining of the cell-scaffold constructs indicated that chondrocyte-like cells were derived from seeded BMSCs and excreted ECM.The cell-scaffold constructs contained pink,smooth and translucent cartilage-like tissue after 3 weeks of culture.We observed evenly distributed cartilage ECM proteoglycans and collagen type Ⅱ around seeded BMSCs on the surface and inside the pores throughout the scaffold.Conclusion This study stuggests that a cartilage ECM scaffold holds much promise for in vitro cartilage tissue engineering.

Stem cells of the reproductive tract of women

Research in stem cells is one of the most rapidly evolving fields of investigation in medicine today. Stem cells are defined as cells that have the capacity to both generate daughter cells identical to the cell of origin (self-renewal) and to produce progeny with more restricted, specialized potential (differentiated cells). This dual ability to self-renew and differentiate offers great promise for expanding our understanding of organ systems, elucidating disease pathophysiology, and creating therapeutic approaches to difficult diseases. The goal of this review is to offer an overview of the different types of stem cells and to provide an introduction to the applications of stem cells to the field of obstetrics and gynecology.

基于“筋、节、骨、髓”分型探讨神经根型颈椎病的手法治疗

神经根型颈椎病(CSR)是一种由于颈椎间盘和椎间关节退变引起的疾病,主要表现为颈部、肩部及上肢神经支配区域的疼痛、感觉异常和运动障碍。传统中医的筋骨辨证在治疗该病时存在概念不清、辨证分型不一等问题。岐黄学者林定坤教授在筋骨辨证的基础上,结合现代解剖学提出了颈椎“筋”“节”“骨”“髓”4大分型。该文通过总结林定坤教授对CSR筋骨辨证学术思想,总结手法治疗该病的作用靶点,提出基于筋劳、节错、骨变和髓损等4种病损分型,采用理筋、调节、护髓和治骨4种手法治疗,以期完善骨伤科手法治疗CSR的辨证施治体系,为临床提供参考。