Selective Cell Retention Technology Research Articles

Autologous Mesenchymal Stromal Cells Combined with Gelatin Sponge for Repair Intervertebral Disc Defect after Discectomy: A Preclinical Study in a Goat Model.

The defect of intervertebral disc (IVD) after discectomy may impair tissue healing and predispose patients to subsequent IVD degeneration, which is thought to be an important cause of recurrence. Cell-based approaches for the treatment of IVD degeneration have shown promise in preclinical studies. However, most of these therapies have not been approved for clinical use due to the risks of abnormal differentiation and microorganism contamination of the culture-expanded cells. Selective cell retention (SCR) technology is non-cultivation technique, which can avoid those preambles in cell expansion. In this study, we used a commercially available BONE GROWTH PROMOTER device (BGP, FUWOSI, Chongqing, China) to concentrate mesenchymal stromal cells (MSCs) from bone marrow aspirate (BMA) through SCR technology. A small incision was made on the L2/3, L3/4 and L4/5 discs of goats and part of nucleus pulposus (NP) was removed to construct IVD defect model. The L2/3 disc was subjected to discectomy only (DO group), the L3/4 disc was implanted with enriched BMA-matrix (CE group), and the L4/5 disc was implanted cultured autologous bone marrow MSCs matrix (CC group). And the intact L1/2 disc served as a non-injured control (NC group). The animals were followed up for 24 weeks after operation. Spine imaging was analysis performed at 4 and 24 weeks. Histology, immunohistochemistry, gene expression and biomechanical analysis were performed to investigate the IVD morphology, content and mechanical properties at 24 weeks. The CE and CC groups showed a significantly smaller reduction in the disc height and T2-weighted signal intensity, and a better spinal segmental stability than DO group. Histological analysis demonstrated that CE and CC groups maintained a relatively well-preserved structure compared to the DO group. Furthermore, real-time PCR and immunohistochemistry demonstrated that aggrecan and type II collagen were up-regulated in CE and CC groups compared to DO group. The strategy of MSCs enrichment combined with gelatin sponge by SCR technology provides a rapid, simple, and effective method for cell concentration and cell-carrier combination. This reparative strategy can be used in clinical treatment of IVD defect after discectomy. NCT03002207.

Laminin alpha 4 promotes bone regeneration by facilitating cell adhesion and vascularization

Selective cell retention (SCR) has been widely used as a bone tissue engineering technique for the real-time fabrication of bone grafts. The greater the number of mesenchymal stem cells (MSCs) and endothelial progenitor cells (EPCs) retained in the scaffold, the better the osteoinductive and angiogenic properties of the scaffold's microenvironment. Improved bioscaffold properties in turn lead to improved bone graft survival, bone regeneration, and angiogenesis. Laminin plays a key role in cell-matrix adhesion, cell proliferation, and differentiation. We designed a collagen-binding domain (CBD) containing the core functional amino acid sequences of laminin α4 (CBD-LN peptide) to supplement the functional surface of a collagen-based decalcified bone matrix (DBM) scaffold. This scaffold promoted MSCs and EPCs early cell adhesion through up-regulating the expression of integrin α5β1 and integrin αvβ3 respectively, thus accelerated the following cell spreading, proliferation, and differentiation. Interestingly, it promoted the retention of MSCs (CD90+/CD105+ cells) and EPCs (CD31+ cells) in the scaffold following the use of clinical SCR technology. Furthermore, the DBM/CBD-LN scaffold induced the formation of type H vessels through the activation of the HIF-1α signaling pathway. The DBM/CBD-LN scaffold displayed rapid bone formation and angiogenesis in vivo, suggesting that it might be used as a new biomaterial in bone tissue engineering. Statement of significanceSelective cell retention technology (SCR) has been utilized in clinical settings to manufacture bioactive bone grafts. Specifically, demineralized bone matrix (DBM) is a widely-used SCR clinical biomaterial but it displays poor adhesion performance and angiogenic activity. In this work, we designed a collagen-binding domain (CBD) containing the core functional amino acid sequences of laminin α4 to supplement the functional surface of a collagen-based DBM scaffold. This bioscaffold promoted SCR-mediated MSCs and EPCs early cell adhesion, thus accelerated the following cell spreading, proliferation, and differentiation. Our results indicate this bioscaffold greatly induced osteogenesis and angiogenesis in vivo. In general, this bioscaffold has a good prospect for SCR application and may provide highly bioactive bone implant in clinical environment.

The clinical use of the enriched bone marrow obtained by selective cell retention technology in treating adolescent idiopathic scoliosis.

PurposeThis retrospective study aimed to evaluate the curative effect of allografts in combination with bone marrow enrichment realised by selective cell retention (SCR) technology in treating adolescent idiopathic scoliosis (AIS).MethodsFrom July 2014 to September 2016, 18 consecutive patients with AIS were treated by posterior fusion and pedicle screw instrumentation. Bone marrow aspirates were obtained and enriched by SCR technology to fabricate bone grafts in combination with allogeneic bones, which were implanted for spinal fusion. Postoperatively, the patients were observed for a minimum of 18 months, with a mean follow-up period of 48 months. The results were assessed both clinically and radiographically. All adverse events and complications were recorded.ResultsA total of 9 male and 9 female patients were included, with an average age of 15.6 years (range, 12–20). The average preoperative Cobb angle was 56° (range, 47°–85°). The average number of levels fused was 11 (range, 9–13). SCR could be accomplished intraoperatively, only consuming approximately 20 ​min. The enriching multiples of measured cellular elements were approximately 2.3–4.2. At final follow-up, the average Cobb angle correction was 83% (range, 61–96%). There was no obvious loss in correction with an average loss of 1.1° (2%). The visual analogue scale score and the Oswestry Disability Index score at final follow-up were significantly ameliorated than those preoperatively. The Scoliosis Research Society 30 questionnaire revealed remarkable improvement in the domains “pain”, “self-image/appearance”, and “satisfaction with management”. There was neither pseudarthrosis nor severe complication.ConclusionThe use of SCR technology could be considered as an effective method for promoting spinal fusion in treating AIS. We proposed a safe, simple, and rapid approach to obtain effective bone grafts for spinal fusion.The translational potential of this articleEnriched bone marrow obtained by selective cell retention technology has the potential to promote spinal fusion for the treatment of adolescent idiopathic scoliosis.

Inhibition of protein-tyrosine phosphatase 1B phosphorylation enhances early adhesion of mesenchymal stem cells to facilitate fabrication of tissue-engineered bone.

Enhancement of cell-matrix adhesion is preferable and crucial in various fields of tissue engineering. Integrins are important receptors that facilitate cell-matrix adhesion, mediated by intracellular molecules and crosstalk with the cadherin adhesion pathway, which mainly facilitates cell-cell adhesion. Protein-tyrosine phosphatase 1B (PTP1B) has emerged as a pivot in the crosstalk between the cadherin adhesion pathway and the integrin adhesion pathway. The phosphorylation state of PTP1B tyrosine-152 (Y152) plays a central role in balancing the two different cell adhesion forms. In this study, a PTP1B Y152 region mimicking (152RM) peptide was designed to decrease the phosphorylation of PTP1B Y152 via competitive inhibition. As a result, the dissociation of cadherin complexes and the release of PTP1B from cadherin had sharply increased, and Src, an important intracellular component of integrin, was activated, indicating that the cadherin adhesion pathway was inhibited, whereas the integrin adhesion pathway was enhanced. Moreover, upon treatment with the 152RM peptide, we observed that the early adhesion of human bone marrow-derived mesenchymal stem cells (MSCs) was accelerated and the anchoring of MSCs on the surface of integrin ligands was enhanced by an enhanced matrix adhesion ability of MSCs themselves. Importantly, the 152RM peptide significantly promoted the adhesion efficiency of MSCs in the selective cell retention technology, which fabricates instant bone implants in clinical settings, to stimulate osteogenesis in vivo.

Multiple integrin ligands provide a highly adhesive and osteoinductive surface that improves selective cell retention technology

Among various bone tissue engineering strategies, selective cell retention (SCR) technology has been used as a practical clinical method for bone graft manufacturing in real time. The more mesenchymal stem cells (MSCs) are retained, the better the osteoinductive microenvironment provided by the scaffold, which in turn promotes the osteogenesis of the SCR-fabricated bone grafts. Integrin receptors are crucial to cell-matrix adhesion and signal transduction. We designed a collagen-binding domain (CBD)-containing IKVAV-cRGD peptide (CBD-IKVAV-cRGD peptide) to complement the collagen-based demineralized bone matrix (DBM) with a functionalized surface containing multiple integrin ligands, which correspond to the highly expressed integrin subtypes on MSCs. This DBM/CBD-IKVAV-cRGD composite exhibited superior in vitro adhesion capacity to cultured MSCs, as determined by oscillatory cell adhesion assay, centrifugal cell adhesion assay and mimetic SCR. Moreover, it promoted the retention of MSC-like CD271+ cells and MSC-like CD90+/CD105+ cells in the clinical SCR method. Furthermore, the DBM/CBD-IKVAV-cRGD composite induced robust MSC osteogenesis, coupled with the activation of the downstream FAK-ERK1/2 signaling pathway of integrins. The SCR-prepared DBM/CBD-IKVAV-cRGD composite displayed superior in vivo osteogenesis, indicating that it may be potentially utilized as a biomaterial in SCR-mediated bone transplantation. Statement of SignificanceSelective cell retention technology (SCR) has been utilized in clinical settings to manufacture bioactive bone grafts. Specifically, demineralized bone matrix (DBM) is a widely-used SCR clinical biomaterial but it displays poor adhesion performance and osteoinduction. Improvements of the DBM that promote cell adhesion and osteoinduction will benefit SCR-prepared implants. In this work, we developed a novel peptide that complements the DBM with a functionalized surface of multiple integrin ligands, which are corresponding to integrin subtypes available on human bone marrow-derived mesenchymal stem cells (MSCs). Our results indicate this novel functionalized bioscaffold greatly increases SCR-mediated MSC adhesion and in vivo osteogenesis. Overall, this novel material has promising SCR applications and may likely provide highly bioactive bone implants in clinical settings.

Long-term Radiological and Clinical Outcomes After Using Bone Marrow Mesenchymal Stem Cells Concentrate Obtained With Selective Retention Cell Technology in Posterolateral Spinal Fusion.

Retrospective study. The aim of this study was to evaluate the long-term clinical and radiological outcomes of the use of bone marrow mesenchymal stem cell concentrate obtained with selective cell retention technology using Cellect with a particular collagen scaffold, Healos for posterolateral spinal fusion. With the increasing rate of spinal fusion, the problem of pseudarthrosis, which contributes to recurrent pain with patient disability, is considered to be the most common cause of revision lumbar spine surgery. Intensive research is being carried out to develop an alternative source of bone grafting and improve the spinal fusion rate. A retrospective review of hospital records was performed. Identified patients were contacted to have a clinical and radiological evaluation follow-up. Clinical outcome was evaluated using visual analog scales for the back pain (VAS), Oswestry Disability Index (ODI) scores, and quality of life (EQ-5D) questionnaire. Radiological outcome was evaluated by performing dynamic flexion/extension lateral views and calculation of segmental Cobb angle. Any implant-associated complication was reported. Computed tomography (CT) scans were also performed. Twenty-one patients were included and all patients achieved successful fusion. The mean difference of the segmental Cobb angle was 0.48° (range 0.3°-0.7°). Computed tomography scans showed solid bilateral fusion with bridging bone (Grade I) in all patients, but solid unilateral fusion with bridging bone (Grade II) was detected for one patient at one level. Patients started to resume working activities within a mean period of 3.5 months. The VAS score for the residual back pain was 4.1 ± 2.1, whereas the ODI was 10.5 ± 5.6 points, and the mean disability index was 21.1%. The use of bone marrow mesenchymal stem cell concentrate obtained with selective cell retention technology could be considered as an effective means for augmenting spinal fusion. 3.

A High-Adhesive Lysine-Cyclic RGD Peptide Designed for Selective Cell Retention Technology.

Cell adhesion is an important property of biomaterials used in selective cell retention (SCR) technology, which fabricates bone grafts rapidly in clinical settings. This could be improved by physical and biologic manipulations. To facilitate retention of the cells on the scaffold, especially osteoprogenitors from bone marrow in the convenient SCR procedure, a lysine-cyclic RGD (LcRGD) peptide was here designed to coordinate positively charged amino acids and the RGD sequence to enhance the adhesion performance of the scaffold. Demineralized bone matrix (DBM) is an important therapeutic resource, but its cell adhesion ability and osteoinductive capacity are low because of its processing. These capabilities can be increased to enhance the performance of DBM when used in SCR technology. Here, LcRGD peptide was used to modify DBM and produce a DBM/LcRGD composite. This composite exhibited enhanced adhesion performance on cultured human bone marrow-derived mesenchymal stem cells and retained more osteoprogenitors from bone marrow than other materials did. The DBM/LcRGD composite displayed a preferable osteoinduction in vitro and osteogenic capacity in vivo. Thus, LcRGD peptide as a commendable modifier of DBM applied in SCR technology can improve bone transplantation.

Osteogenic ability of bone marrow stem cells intraoperatively enriched by a novel matrix.

Poly-L-lysine (PLL) is commonly used as an adhibiting agent due to its good viscosity, and demineralized bone matrix (DBM) is a common enriched matrix for selective cell retention technology. Therefore, the aim of this study was to use PLL to coat the surface and interspaces of DBM to form a novel type of enriched matrix [DBM coated with PLL (PLL-DBM)], in order to effectively improve the enrichment effects of bone marrow stem cells and enhance their osteogenic ability. Electron microscope scanning and the infrared spectrum were used to observe the structure of PLL-DBM and the optimal conditions for the combination of PLL and DBM. Enriching effects on bone marrow nucleated cells (NCs) and platelets (PLTs) were detected with an automated hematology analyzer. The osteogenesis of the following four groups was assessed with a grafting bone model in a goat spinal transverse process: IA, tissue engineered bone (TEB) fabricated following enrichment of bone marrow with PLL-DBM; IB, autogenous iliac bone; IIC, TEB fabricated following enrichment of bone marrow with DBM; IID, blank DBM. The goats were sacrificed in one batch at week 16 after the surgery and the fusion specimens were examined using X-ray and three-dimensional computed tomography (CT). In addition, the CT value was determined and the histology and biomechanics were analyzed in order to evaluate the osteogenic ability. The results showed that PLL and DBM combined well and that PLL-DBM exhibited a natural mesh pore structure. The fold enrichment of NCs and PLTs with PLL-DBM was significantly higher than that with DBM. The fusion effects of the IA and IB groups were similar and significantly enhanced compared with those of the IIC and IID groups. The results confirmed that PLL-DBM is an ideal enriched matrix for bone marrow stem cells, and TEB rapidly fabricated by PLL-DBM intraoperatively enriched bone marrow stem cells exhibits an improved osteogenic ability.

A composite demineralized bone matrix – Self assembling peptide scaffold for enhancing cell and growth factor activity in bone marrow

The need for suitable bone grafts is high; however, there are limitations to all current graft sources, such as limited availability, the invasive harvest procedure, insufficient osteoinductive properties, poor biocompatibility, ethical problems, and degradation properties. The lack of osteoinductive properties is a common problem. As an allogenic bone graft, demineralized bone matrix (DBM) can overcome issues such as limited sources and comorbidities caused by invasive harvest; however, DBM is not sufficiently osteoinductive. Bone marrow has been known to magnify osteoinductive components for bone reconstruction because it contains osteogenic cells and factors. Mesenchymal stem cells (MSCs) derived from bone marrow are the gold standard for cell seeding in tissue-engineered biomaterials for bone repair, and these cells have demonstrated beneficial effects. However, the associated high cost and the complicated procedures limit the use of tissue-engineered bone constructs. To easily enrich more osteogenic cells and factors to DBM by selective cell retention technology, DBM is modified by a nanoscale self-assembling peptide (SAP) to form a composite DBM/SAP scaffold. By decreasing the pore size and increasing the charge interaction, DBM/SAP scaffolds possess a much higher enriching yield for osteogenic cells and factors compared with DBM alone scaffolds. At the same time, SAP can build a cellular microenvironment for cell adhesion, proliferation, and differentiation that promotes bone reconstruction. As a result, a suitable bone graft fabricated by DBM/SAP scaffolds and bone marrow represents a new strategy and product for bone transplantation in the clinic.

Efficacy of Mesenchymal Stem Cell Enriched Grafts in an Ovine Posterolateral Lumbar Spine Model

Four groups of 6 animals underwent single-level noninstrumented posterolateral lumbar fusion (PLF) with one of the following grafts: 1) autograft, 2) cell-enriched beta-tricalcium phosphate (TCP), 3) TCP with whole bone marrow, and 4) TCP alone. Plain radiographs were taken after surgery and at death, 6 months after surgery. Explanted spine segments were analyzed by manual palpation, micro-CT, and histology. A sheep spine fusion study was undertaken to evaluate the healing performance of a TCP graft enriched with osteoprogenitor cells using Selective Cell Retention technology (SCR), compared with autograft, TCP with whole bone marrow, and TCP alone. Improved bone healing with previously demonstrated using grafts enriched in osteoprogenitor cells. Cell-enriched grafts were obtained by processing 30 mL of bone marrow through 10 mL of TCP. TCP was also used either saturated with bone marrow or alone. At 6 months, 33% of the SCR-enriched TCP and 25% of the autograft sites were fused, compared with 8% of the TCP plus whole bone marrow and 0% of the TCP alone. Histology of fused samples showed denser bone formation in the SCR-enriched TCP grafts than in the autograft sites. The use of SCR-enriched TCP and autograft resulted in similar fusion rates in an ovine posterolateral noninstrumented lumbar spine fusion model.

Bone grafts prepared with selective cell retention technology heal canine segmental defects as effectively as autograft

Using a canine critical-size segmental defect model, a two-phased study was undertaken to evaluate the healing efficacy of demineralized bone and cancellous chips (DBM-CC) enriched with osteoprogenitor cells using a Selective Cell Retention (SCR) technology. The goals of this study were: 1) to determine the bone-healing efficacy of SCR-enriched grafts versus autograft, and 2) to assess the value of clotting SCR-enriched grafts with platelet-rich plasma (PRP). Thirty dogs were included in Phase I: 18 dogs were treated with an SCR-enriched DBM-CC graft clotted with autologous bone marrow, and were compared to 12 autograft controls. In Phase II, 24 animals were divided into 4 groups of 6 animals, each treated with a different bone graft material: 1) iliac crest autograft, 2) DBM-CC alone, 3) DBM-CC saturated with marrow, and 4) SCR-enriched DBM-CC clotted with PRP. All grafts were placed unilaterally in a 21-mm long osteoperiosteal femoral, instrumented, critical-size defect. Radiographs were obtained for all animals postoperatively and every 4-16 weeks; animals were then sacrificed. All femurs were prepared for histology. Femurs in the Phase II study were also analyzed by micro-CT. At 16 weeks, healing--defined by bridging bone across the defects--was observed in 50% of the DBM-CC alone group and 67% of the DBM-CC saturated with marrow group; 100% of the autograft and SCR-enriched DBM-CC groups were healed. Histologically, grafts clotted with PRP showed more mature bone than those implanted with autologous bone, which in turn were similar to those implanted with bone marrow clotted SCR-enriched grafts. These results demonstrated that: 1) SCR-enriched DBM-CC was equivalent to autograft to repair critical-size defects, and 2) while not statistically significant, PRP may have accelerated bone maturation when used to clot osteoprogenitor-enriched DBM-CC grafts--as compared to cell-enriched, DBM-CC grafts without PRP--in large animal models.

98. Selective cell retention technology in spinal fusion

98. Selective cell retention technology in spinal fusion