In regenerative medicine,the isolation of mesenchymal stromal cells(MSCs)from the adipose tissue’s stromal vascular fraction(SVF)is a critical area of study.Our review meticulously examines the isolation process of M...In regenerative medicine,the isolation of mesenchymal stromal cells(MSCs)from the adipose tissue’s stromal vascular fraction(SVF)is a critical area of study.Our review meticulously examines the isolation process of MSCs,starting with the extraction of adipose tissue.The choice of liposuction technique,anatomical site,and immediate processing are essential to maintain cell functionality.We delve into the intricacies of enzymatic digestion,emphasizing the fine-tuning of enzyme concentrations to maximize cell yield while preventing harm.The review then outlines the filtration and centrifugation techniques necessary for isolating a purified SVF,alongside cell viability assessments like flow cytometry,which are vital for confirming the efficacy of the isolated MSCs.We discuss the advantages and drawbacks of using autologous vs allogeneic SVF sources,touching upon immunocompatibility and logistical considerations,as well as the variability inherent in donor-derived cells.Anesthesia choices,the selection between hypo-dermic needles vs liposuction cannulas,and the role of adipose tissue lysers in achieving cellular dissociation are evaluated for their impact on SVF isolation.Centrifugation protocols are also analyzed for their part in ensuring the integrity of the SVF.The necessity for standardized MSC isolation protocols is highlighted,promoting reproducibility and successful clinical application.We encourage ongoing research to deepen the understanding of MSC biology and therapeutic action,aiming to further the field of regenerative medicine.The review concludes with a call for rigorous research,interdisciplinary collaboration,and strict adherence to ethical and regulatory standards to safeguard patient safety and optimize treatment outcomes with MSCs.展开更多
BACKGROUND Current osteoarthritis(OA)treatments focus on symptom relief without addressing the underlying disease process.In regenerative medicine,current treatments have limitations.In regenerative medicine,more rese...BACKGROUND Current osteoarthritis(OA)treatments focus on symptom relief without addressing the underlying disease process.In regenerative medicine,current treatments have limitations.In regenerative medicine,more research is needed for intra-articular stromal vascular fraction(SVF)injections in OA,including dosage optimization,long-term efficacy,safety,comparisons with other treatments,and mechanism exploration.AIM To compare the efficacy of intra-articular SVF with corticosteroid(ICS)injections in patients with primary knee OA.METHODS The study included 50 patients with Kellgren-Lawrence grades II and III OA.Patients were randomly assigned(1:1)to receive either a single intra-articular SVF injection(group A)or a single intra-articular ICS(triamcinolone)(group B)injection.Patients were followed up at 1,3,6,12,and 24 months.Visual analog score(VAS)and International Knee Documentation Committee(IKDC)scores were administered before the procedure and at all followups.The safety of SVF in terms of adverse and severe adverse events was recorded.Statistical analysis was performed with SPSS Version 26.0,IBM Corp,Chicago,IL,United States.RESULTS Both groups had similar demographics and baseline clinical characteristics.Follow-up showed minor patient loss,resulting in 23 and 24 in groups A and B respectively.Group A experienced a notable reduction in pain,with VAS scores decreasing from 7.7 to 2.4 over 24 months,compared to a minor reduction from 7.8 to 6.2 in Group B.This difference in pain reduction in group A was statistically significant from the third month onwards.Additionally,Group A showed significant improvements in knee functionality,with IKDC scores rising from 33.4 to 83.10,whereas Group B saw a modest increase from 36.7 to 45.16.The improvement in Group A was statistically significant from 6 months and maintained through 24 months.CONCLUSION Our study demonstrated that intra-articular administration of SVF can lead to reduced pain and improved knee function in patients with primary knee OA.More adequately powered,multi-center,double-blinded,randomised clinical trials with longer follow-ups are needed to further establish safety and justify its clinical use.展开更多
Background:The stromal vascular fraction(SVF),a cluster of stem and progenitor cells isolated from adipose tissue,holds significant promise for application in regenerative medicine.However,the existing methods for SVF...Background:The stromal vascular fraction(SVF),a cluster of stem and progenitor cells isolated from adipose tissue,holds significant promise for application in regenerative medicine.However,the existing methods for SVF isolation are time-consuming and expensive.Thus,in this study,we explored a new method of SVF extrac-tion-ultrasound-assisted SVF isolation(USASI)-and compared the viability and characteristics of SVF isolated using different methods.Methods:SVF extraction methods using different combinations of ultrasound power,ultrasound time,collagenase dosage,and collagenase digestion time were compared with those of the control group(collagenase digestion method).The cell yield and vitality of the SVF were evaluated via cell counting and trypan blue staining.The cell components and immunophenotypes of freshly isolated SVF were analyzed using flow cytometry.The prolifer-ative capacity and differentiation potential of the SVF were also identified.Results:Ultrasonication at 95 W-20 kHz for 30 s followed by digestion with 0.15%collagenase for 30 min was identified as the most suitable parameter for the USASI method in isolating SVF,as recommended based on the evaluation of various tested conditions.The USASI method significantly reduced the collagenase dosage and shortened the digestion time.Compared to the collagenase digestion method,the USASI method had a higher cell yield and cell viability,with no adverse effects on cell components,proliferative capacity,or multipotential differentiation capacity.Conclusion:With reduced processing time,lower collagenase dosage,and increased cell yield without impairing the viability and characteristics of SVF,USASI holds the potential to emerge as a time-saving and cost-effective method for future clinical applications.展开更多
Recently,we read a mini-review published by Jeyaraman et al.The article explored the optimal methods for isolating mesenchymal stromal cells from adipose tissue-derived stromal vascular fraction(SVF).Key factors inclu...Recently,we read a mini-review published by Jeyaraman et al.The article explored the optimal methods for isolating mesenchymal stromal cells from adipose tissue-derived stromal vascular fraction(SVF).Key factors include tissue source,processing techniques,cell viability assessment,and the advantages/disadvantages of autologous vs allogeneic use.The authors emphasized the need for standardized protocols for SVF isolation,ethical and regulatory standards for cell-based therapy,and safety to advance mesenchymal stromal cell-based therapies in human patients.This manuscript shares our perspective on SVF isolation in canines.We discussed future directions to potentiate effective regenerative medicine therapeutics in human and veterinary medicine.展开更多
BACKGROUND Adipose-derived stem cells(ADSCs)and the stromal vascular fraction(SVF)have garnered substantial interest in regenerative medicine due to their potential to treat a wide range of conditions.Traditional enzy...BACKGROUND Adipose-derived stem cells(ADSCs)and the stromal vascular fraction(SVF)have garnered substantial interest in regenerative medicine due to their potential to treat a wide range of conditions.Traditional enzymatic methods for isolating these cells face challenges such as high costs,lengthy processing time,and regulatory complexities.AIM This systematic review aimed to assess the efficacy and practicality of nonenzymatic,mechanical methods for isolating SVF and ADSCs,comparing these to conventional enzymatic approaches.METHODS Following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines,a comprehensive literature search was conducted across multiple databases.Studies were selected based on inclusion criteria focused on non-enzymatic isolation methods for SVF and ADSCs from adipose tissue.The risk of bias was assessed,and a qualitative synthesis of findings was performed due to the methodological heterogeneity of the included studies.RESULTS Nineteen studies met the inclusion criteria,highlighting various mechanical techniques such as centrifugation,vortexing,and ultrasonic cavitation.The review identified significant variability in cell yield and viability,and the integrity of isolated cells across different non-enzymatic methods compared to enzymatic procedures.Despite some advantages of mechanical methods,including reduced processing time and avoidance of enzymatic reagents,the evidence suggests a need for optimization to match the cell quality and therapeutic efficacy achievable with enzymatic isolation.CONCLUSION Non-enzymatic,mechanical methods offer a promising alternative to enzymatic isolation of SVF and ADSCs,potentially simplifying the isolation process and reducing regulatory hurdles.However,further research is necessary to standardize these techniques and ensure consistent,high-quality cell yields for clinical applications.The development of efficient,safe,and reproducible non-enzymatic isolation methods could significantly advance the field of regenerative medicine.展开更多
Background: Regenerative medicine holds promise for treating degenerative and inflammatory conditions like osteoarthritis (OA). However, the complex molecular mechanisms of OA and the limitations of current therapies ...Background: Regenerative medicine holds promise for treating degenerative and inflammatory conditions like osteoarthritis (OA). However, the complex molecular mechanisms of OA and the limitations of current therapies remain challenges. Adipose-derived stem cells (ADSC) and stromal vascular fraction (SVF) are gaining attention for OA treatment due to their abundance in adipose tissue. The literature delineates two primary techniques for SVF extraction from adipose tissue: enzymatic digestion and mechanical methods. The Lipocube Hybrid SVF presents a straightforward and secure mechanical isolation method for SVF, enhancing its regenerative potential for various applications. Purpose: This study aims to provide valuable insights into the potential of Lipocube Hybrid SVF as a regenerative therapy for OA, contributing to the broader understanding of its applicability in addressing this debilitating condition. Method: To assess the effectiveness and safety of the Lipocube Hybrid SVF, we have designed a comparative study that evaluates cellular activity and viability, phenotypic characterization, and differentiation potential. The in vitro activity of mechanically isolated SVF is compared to the established gold standard enzymatic digestion method. After in vitro studies, Lipocube Hybrid mechanical isolation method was used to isolate SVF and applied in 42 knee and 7 hip joints of 28 patients with Grade II, Grade III, and Grade IV OA. Results: The Lipocube Hybrid group had slightly lower viable cell numbers but higher cell viability. Flow cytometry analysis showed the Lipocube Hybrid group exhibited more favorable markers for regenerative potential and reduced inflammatory response. Additionally, both groups demonstrated successful osteogenic differentiation, with the Lipocube Hybrid group excelling in chondrogenic and adipogenic differentiation. The clinical application of the Lipocube Hybrid SVF in OA patients resulted in significant improvements in WOMAC and VAS scores across different OA grades. Conclusions: This comparative study was conducted to evaluate the effectiveness and safety of the Lipocube Hybrid SVF, which has shown promise in laboratory settings, for different stages of osteoarthritis. The study findings provide valuable insights into the potential of Lipocube Hybrid SVF as a regenerative therapy for OA, highlighting its suitability for addressing this debilitating condition.展开更多
AIM: To investigate the role of vascular endothelial growth factor (VEGF/and its receptors VEGFR-1 and 2 in the growth and differentiation of gastrointestinal strornal tumors (GISTs). METHODS: Thirty-three GISTs,...AIM: To investigate the role of vascular endothelial growth factor (VEGF/and its receptors VEGFR-1 and 2 in the growth and differentiation of gastrointestinal strornal tumors (GISTs). METHODS: Thirty-three GISTs, 15 leiomyomas and 6 schwannomas were examined by immunohistochemistry in this study. RESULTS: VEGF protein was expressed in the cytoplasm of tumor cells, and VEGFRol and 2 were expressed both in the cytoplasm and on the membrane of all tumors. Irnrnunohistochernical staining revealed that 26 GISTs (78.8%), 9 leiornyornas (60.0%) and 3 schwannornas (50.0%/were positive for VEGF; 24 GISTs (72.7%/, 12 leiornyornas (80.0%) and 4 schwannornas (66.7%) were positive for VEGFR-1; 30 GISTs (90.9%/, 5 leiornyornas (33.3%/and 4 schwannornas (66.7%) were positive for VEGFR-2. VEGFR-2 expression was statistically different between GISTs and leiomyomas (P 〈 0.0001). However, there was no correlation between the expression of VEGF pathway componenets and the clinical risk categories. CONCLUSION: Our results suggest that the VEGF pathway may play an important role in the differentiation of GISTs, leiomyomas and schwannomas.展开更多
AIM: To estimate whether S-TI571 inhibits the expression of vascular endothelial growth factor (VEGF) in the gastrointestinal stromal tumor (GIST) cells. METHODS: We used GIST cell line, GIST-T1. It has a hetero...AIM: To estimate whether S-TI571 inhibits the expression of vascular endothelial growth factor (VEGF) in the gastrointestinal stromal tumor (GIST) cells. METHODS: We used GIST cell line, GIST-T1. It has a heterogenic 57-bp deletion in exon 11 to produce a mutated c-KIT, which results in constitutive activation of c-KIT. Cells were treated with/without STI571 or stem cell factor (SCF). Transcription and expression of VEGF were determined by RT-PCR and flow cytometry or Western blotting, respectively. Activated c-KIT was estimated by immunoprecipitation analysis. Cell viability was determined by PITT assay. RESULTS: Activation of c-KIT was inhibited by STI571 treatment. VEGF was suppressed at both the transcriptional and translational levels in a temporal and dose-dependent manner by STI571. SCF upregulated the expression of VEGF and it was inhibited by S-13571. STI571 also reduced the cell viability of the GIST-T1 cells, as determined by PTT assay. CONCLUSION: Activation of c-KIT in the GIST-T1 regulated the expression of VEGF and it was inhibited by ST571. STI571 has antitumor effects on the GIST cells with respect to not only the inhibition of cell growth, but also the suppression of VEGF expression.展开更多
BACKGROUND The development of fully functional small diameter vascular grafts requires both a properly defined vessel conduit and tissue-specific cellular populations.Mesenchymal stromal cells(MSCs) derived from the W...BACKGROUND The development of fully functional small diameter vascular grafts requires both a properly defined vessel conduit and tissue-specific cellular populations.Mesenchymal stromal cells(MSCs) derived from the Wharton's Jelly(WJ) tissue can be used as a source for obtaining vascular smooth muscle cells(VSMCs),while the human umbilical arteries(h UAs) can serve as a scaffold for blood vessel engineering.AIM To develop VSMCs from WJ-MSCs utilizing umbilical cord blood platelet lysate.METHODS WJ-MSCs were isolated and expanded until passage(P) 4. WJ-MSCs were properly defined according to the criteria of the International Society for Cell and Gene Therapy. Then, these cells were differentiated into VSMCs with the use of platelet lysate from umbilical cord blood in combination with ascorbic acid,followed by evaluation at the gene and protein levels. Specifically, gene expression profile analysis of VSMCs for ACTA2, MYH11, TGLN, MYOCD, SOX9,NANOG homeobox, OCT4 and GAPDH, was performed. In addition,immunofluorescence against ACTA2 and MYH11 in combination with DAPI staining was also performed in VSMCs. HUAs were decellularized and served as scaffolds for possible repopulation by VSMCs. Histological and biochemical analyses were performed in repopulated h UAs.RESULTS WJ-MSCs exhibited fibroblastic morphology, successfully differentiating into"osteocytes", "adipocytes" and "chondrocytes", and were characterized by positive expression(> 90%) of CD90, CD73 and CD105. In addition, WJ-MSCs were successfully differentiated into VSMCs with the proposed differentiation protocol. VSMCs successfully expressed ACTA2, MYH11, MYOCD, TGLN and SOX9. Immunofluorescence results indicated the expression of ACTA2 and MYH11 in VSMCs. In order to determine the functionality of VSMCs, h UAs were isolated and decellularized. Based on histological analysis, decellularized h UAs were free of any cellular or nuclear materials, while their extracellular matrix retained intact. Then, repopulation of decellularized h UAs with VSMCs was performed for 3 wk. Decellularized h UAs were repopulated efficiently by the VSMCs. Biochemical analysis revealed the increase of total hydroyproline and s GAG contents in repopulated h UAs with VSMCs. Specifically, total hydroxyproline and s GAG content after the 1 st, 2 nd and 3 rd wk was 71 ± 10, 74 ± 9 and 86 ± 8 μg hydroxyproline/mg of dry tissue weight and 2 ± 1, 3 ± 1 and 3 ± 1μg s GAG/mg of dry tissue weight, respectively. Statistically significant differences were observed between all study groups(P<0.05).CONCLUSION VSMCs were successfully obtained from WJ-MSCs with the proposed differentiation protocol. Furthermore, h UAs were efficiently repopulated by VSMCs. Differentiated VSMCs from WJ-MSCs could provide an alternative source of cells for vascular tissue engineering.展开更多
Osteoarthritis(OA)is considered to be a highly heterogeneous disease with progressive cartilage loss,subchondral bone remodeling,and low-grade inflammation.It is one of the world's leading causes of disability.Mos...Osteoarthritis(OA)is considered to be a highly heterogeneous disease with progressive cartilage loss,subchondral bone remodeling,and low-grade inflammation.It is one of the world's leading causes of disability.Most conventional clinical treatments for OA are palliative drugs,which cannot fundamentally cure this disease.The stromal vascular fraction(SVF)from adipose tissues is a heterogeneous cell population.According to previous studies,it contains a large number of mesenchymal stem cells,which have been used to treat OA with good therapeutic results.This safe,simple,and effective therapy is expected to be applied and promoted in the future.In this paper,the detailed pathogenesis,diagnosis,and current clinical treatments for OA are introduced.Then,clinical studies and the therapeutic mechanism of SVF for the treatment of OA are summarized.展开更多
AIM:To characterize the implications of vascular endothelial growth factor(VEGF)-A in stromal cells and colorectal cancer and the expression of VEGF-A splice variants.METHODS:VEGF-A expression in tumor and stromal cel...AIM:To characterize the implications of vascular endothelial growth factor(VEGF)-A in stromal cells and colorectal cancer and the expression of VEGF-A splice variants.METHODS:VEGF-A expression in tumor and stromal cells from 165 consecutive patients with colorectal cancer was examined by immunohistochemistry.The association between VEGF-A expression status and clinicopathological factors was investigated.Twenty freshfrozen samples were obtained for laser capture microdissection to analyze the splice variants of VEGF-A.RESULTS:VEGF-A was expressed in 53.9% and 42.4% of tumor and stromal cells,respectively.VEGF-A expression in tumor cells(t-VEGF-A) was associated with advanced clinical stage(stage 0,1/9;stage 1,2/16;stage 2,32/55;stage 3,38/66;stage 4,16/19,P < 0.0001).VEGF-A expression in stromal cells(s-VEGF-A) increased in the earlier clinical stage(stage 0,7/9;stage 1,6/16;stage 2,33/55;stage 3,22/66;stage 4,5/19;P = 0.004).Multivariate analyses for risk factors of recurrence showed that only s-VEGF-A expression was an independent risk factor for recurrence(relative risk 0.309,95% confidence interval 0.141-0.676,P = 0.0033).The five-year disease-free survival(DFS) rates of t-VEGF-A-positive and-negative cases were 51.4% and 62.9%,respectively.There was no significant difference in t-VEGF-A expression status.The five-year DFS rates of s-VEGF-A-positive and-negative cases were 73.8% and 39.9%,respectively.s-VEGFA-positive cases had significantly better survival than s-VEGF-A-negative cases(P = 0.0005).Splice variant analysis revealed that t-VEGF-A was mainly composed of VEGF165 and that s-VEGF-A included both VEGF165 and VEGF165b.In cases with no venous invasion(v0),the level of VEGF165b mRNA was significantly higher(v0 204.5 ± 122.7,v1 32.5 ± 36.7,v2 2.1 ± 1.7,P = 0.03).The microvessel density tended to be lower in cases with higher VEGF165b mRNA levels.CONCLUSION:s-VEGF-A appears be a good prognostic factor for colorectal cancer and includes VEGF165 and VEGF165b.展开更多
BACKGROUND:Studies have demonstrated that bone marrow stromal cells (BMSCs) undergo neuronal differentiation under certain in vitro conditions.However,very few inducers of BMSC differentiation have been used in cli...BACKGROUND:Studies have demonstrated that bone marrow stromal cells (BMSCs) undergo neuronal differentiation under certain in vitro conditions.However,very few inducers of BMSC differentiation have been used in clinical application.The effects of vascular endothelial growth factor (VEGF) on in vitro neuronal differentiation of BMSCs remain poorly understood.OBJECTIVE:To investigate the effect of VEGF on neuronal differentiation of BMSCs in vitro,and to determine the best VEGF concentration for experimental induction.DESIGN,TIME AND SETTING:In vitro comparative study was performed at the Central Laboratory and Laboratory of Male Reproductive Medicine,Shenzhen Hospital of Peking University from October 2008 to August 2009.MATERIALS:Recombinant human VEGF165 was purchased from Peprotech Asia,Rehovot,Israel.Neuron-specific enolase (NSE) was purchased from Beijing Biosynthesis Biotechnology,China.METHODS:BMSCs were harvested from adult Sprague Dawley rats.The passaged cells were pre-induced with 10 ng/mL basic fibroblast growth factor for 24 hours,followed by differentiation induction with 0,5,10,and 20 ng/mL VEGF,respectively.MAIN OUTCOME MEASURES:Morphological changes in BMSCs prior to and following VEGF induction.Expression of NSE following induction was determined by immunocytochemistry.RESULTS:Shrunken,round cells,with a strong refraction and thin bipolar or multipolar primary and secondary branches were observed 3 days after induction with 5,10,and 20 ng/mL VEGF.However,these changes were not observed in the control group.At 10 days after induction,the number of NSE-positive cells was greatest in the 10 ng/mL VEGF-treated group (P〈 0.05).The number of NSE-positive cells was least in the control group at 3 and 10 days post-induction (P〈 0.05).Moreover,the number of NSE-positive cells was greater at 10 days compared with at 3 days after induction (P〈 0.05).CONCLUSION:Of the VEGF concentrations tested,10 ng/mL induced the greatest number of neuronal-like cells in vitro from BMSCs.展开更多
The increasing implementation of multicentre studies has led to a need for the optimization of a method that allows for accurate post-hoc analysis of patient biological samples. Assessment of total cell number, viabil...The increasing implementation of multicentre studies has led to a need for the optimization of a method that allows for accurate post-hoc analysis of patient biological samples. Assessment of total cell number, viability and immunophenotype can present logistical challenges which can be aided by batch processing. The increased sample storage time that this requires necessitates the use of reagents to preserve cellular integrity, viability and immunophenotype. TransFix is a stabilising reagent that has been developed for the preservation of cell numbers and cell marker expression in peripheral whole blood for up to ten days. This study investigated the use of TransFix reagent for the preservation of the stromal vascular fraction (SVF) of collagenase digested adipose tissue. It was demonstrated that TransFix was suitable for accurately measuring nucleated SVF cell numbers for up to seven days as well as back calculating original cell viability. It also stabilised three CD markers commonly used to identify populations within SVF (CD90, CD31 and CD45) for up to seven days. There was no significant difference between the number of CD90, CD31 and CD45 positive cells after stabilisation at Day 7 compared to Day 0 unstabilised samples. The results suggest that TransFix can be used to preserve a biological mixed cell population from human adipose-derived SVF for up to seven days for accurate post-hoc analysis.展开更多
Lipotransfer has become a powerful regenerative tool,largely because of its cellular components,the stromal vascular fraction(SVF).However,the clinical separation of cells with collagenase is strictly legislated.In 20...Lipotransfer has become a powerful regenerative tool,largely because of its cellular components,the stromal vascular fraction(SVF).However,the clinical separation of cells with collagenase is strictly legislated.In 2017,Yao et al.postulated a novel fat-derived product mechanically concentrating SVF cells and an extracellular matrix(ECM)and named it stromal vascular fraction gel(SVF-gel).This review discussed the protocol of SVF-gel and its component as well as its inner structure.The histologic examination and the retention rate after the transplantation of SVF-gel were also rendered.Moreover,we summed up the rejuvenating and regenerative use of SVF-gel and introduced its possible mechanism.展开更多
Objective To investigate the effect of adipose stromal vascular fraction cells(SVFs)on the survival rate of fat ransplantation.Methods 0.5mL autologous fat tissue was mixed with: ① DiI-labeled autologous SVFs (Group ...Objective To investigate the effect of adipose stromal vascular fraction cells(SVFs)on the survival rate of fat ransplantation.Methods 0.5mL autologous fat tissue was mixed with: ① DiI-labeled autologous SVFs (Group A);②展开更多
BACKGROUND Gastrointestinal stromal tumors(GISTs)are considered the most common mesenchymal tumors of the gastrointestinal tract.Microvessel density(MVD)constitutes a direct method of vascularity quantification and ha...BACKGROUND Gastrointestinal stromal tumors(GISTs)are considered the most common mesenchymal tumors of the gastrointestinal tract.Microvessel density(MVD)constitutes a direct method of vascularity quantification and has been associated with survival rates in multiple malignancies.AIM To appraise the effect of MVD on the survival of patients with GIST.METHODS This study adhered to Systematic reviews and Meta-Analyses guidelines and the Cochrane Handbook for Systematic Reviews of Interventions.Electronic scholar databases and grey literature repositories were systematically screened.The Fixed Effects or Random Effects models were used according to the Cochran Q test.RESULTS In total,6 eligible studies were identified.The pooled hazard ratio(HR)for disease free survival(DFS)was 8.52(95%CI:1.69-42.84,P=0.009).The odds ratios of disease-free survival between high and low MVD groups at 12 and 60 mo did not reach statistical significance.Significant superiority of the low MVD group in terms of DFS was documented at 36 and 120 mo(OR:8.46,P<0.0001 and OR:22.71,P=0.0003,respectively)as well as at metastases rate(OR:0.11,P=0.0003).CONCLUSION MVD significantly correlates with the HR of DFS and overall survival rates at 36 and 120 mo.Further prospective studies of higher methodological quality are required.展开更多
文摘In regenerative medicine,the isolation of mesenchymal stromal cells(MSCs)from the adipose tissue’s stromal vascular fraction(SVF)is a critical area of study.Our review meticulously examines the isolation process of MSCs,starting with the extraction of adipose tissue.The choice of liposuction technique,anatomical site,and immediate processing are essential to maintain cell functionality.We delve into the intricacies of enzymatic digestion,emphasizing the fine-tuning of enzyme concentrations to maximize cell yield while preventing harm.The review then outlines the filtration and centrifugation techniques necessary for isolating a purified SVF,alongside cell viability assessments like flow cytometry,which are vital for confirming the efficacy of the isolated MSCs.We discuss the advantages and drawbacks of using autologous vs allogeneic SVF sources,touching upon immunocompatibility and logistical considerations,as well as the variability inherent in donor-derived cells.Anesthesia choices,the selection between hypo-dermic needles vs liposuction cannulas,and the role of adipose tissue lysers in achieving cellular dissociation are evaluated for their impact on SVF isolation.Centrifugation protocols are also analyzed for their part in ensuring the integrity of the SVF.The necessity for standardized MSC isolation protocols is highlighted,promoting reproducibility and successful clinical application.We encourage ongoing research to deepen the understanding of MSC biology and therapeutic action,aiming to further the field of regenerative medicine.The review concludes with a call for rigorous research,interdisciplinary collaboration,and strict adherence to ethical and regulatory standards to safeguard patient safety and optimize treatment outcomes with MSCs.
文摘BACKGROUND Current osteoarthritis(OA)treatments focus on symptom relief without addressing the underlying disease process.In regenerative medicine,current treatments have limitations.In regenerative medicine,more research is needed for intra-articular stromal vascular fraction(SVF)injections in OA,including dosage optimization,long-term efficacy,safety,comparisons with other treatments,and mechanism exploration.AIM To compare the efficacy of intra-articular SVF with corticosteroid(ICS)injections in patients with primary knee OA.METHODS The study included 50 patients with Kellgren-Lawrence grades II and III OA.Patients were randomly assigned(1:1)to receive either a single intra-articular SVF injection(group A)or a single intra-articular ICS(triamcinolone)(group B)injection.Patients were followed up at 1,3,6,12,and 24 months.Visual analog score(VAS)and International Knee Documentation Committee(IKDC)scores were administered before the procedure and at all followups.The safety of SVF in terms of adverse and severe adverse events was recorded.Statistical analysis was performed with SPSS Version 26.0,IBM Corp,Chicago,IL,United States.RESULTS Both groups had similar demographics and baseline clinical characteristics.Follow-up showed minor patient loss,resulting in 23 and 24 in groups A and B respectively.Group A experienced a notable reduction in pain,with VAS scores decreasing from 7.7 to 2.4 over 24 months,compared to a minor reduction from 7.8 to 6.2 in Group B.This difference in pain reduction in group A was statistically significant from the third month onwards.Additionally,Group A showed significant improvements in knee functionality,with IKDC scores rising from 33.4 to 83.10,whereas Group B saw a modest increase from 36.7 to 45.16.The improvement in Group A was statistically significant from 6 months and maintained through 24 months.CONCLUSION Our study demonstrated that intra-articular administration of SVF can lead to reduced pain and improved knee function in patients with primary knee OA.More adequately powered,multi-center,double-blinded,randomised clinical trials with longer follow-ups are needed to further establish safety and justify its clinical use.
基金supported by the National Natural Science Foundation of China(grant nos.81971848 and 82272287)Shanghai Municipal Key Clinical Specialty(grant no,shslczdzk00901)+2 种基金Clinical Research Plan of SHDC(rant nos.SHDC2020CR1019B and SHC2020CR402)Innovative Research Team of High-Level Local Universities in Shanghai(grant no.SSMU-ZDCX20180700)Shanghai Clinical Research Center of Plastic and Reconstructive Surgery supported by the Science and Technology Commission of Shanghai Municipality(grant no.22MC1940300).
文摘Background:The stromal vascular fraction(SVF),a cluster of stem and progenitor cells isolated from adipose tissue,holds significant promise for application in regenerative medicine.However,the existing methods for SVF isolation are time-consuming and expensive.Thus,in this study,we explored a new method of SVF extrac-tion-ultrasound-assisted SVF isolation(USASI)-and compared the viability and characteristics of SVF isolated using different methods.Methods:SVF extraction methods using different combinations of ultrasound power,ultrasound time,collagenase dosage,and collagenase digestion time were compared with those of the control group(collagenase digestion method).The cell yield and vitality of the SVF were evaluated via cell counting and trypan blue staining.The cell components and immunophenotypes of freshly isolated SVF were analyzed using flow cytometry.The prolifer-ative capacity and differentiation potential of the SVF were also identified.Results:Ultrasonication at 95 W-20 kHz for 30 s followed by digestion with 0.15%collagenase for 30 min was identified as the most suitable parameter for the USASI method in isolating SVF,as recommended based on the evaluation of various tested conditions.The USASI method significantly reduced the collagenase dosage and shortened the digestion time.Compared to the collagenase digestion method,the USASI method had a higher cell yield and cell viability,with no adverse effects on cell components,proliferative capacity,or multipotential differentiation capacity.Conclusion:With reduced processing time,lower collagenase dosage,and increased cell yield without impairing the viability and characteristics of SVF,USASI holds the potential to emerge as a time-saving and cost-effective method for future clinical applications.
基金Supported by the Department of Biotechnology,Ministry of Science and Technology,Government of India,New Delhi,No.BT/PR42179/AAQ/1/814/2021SERB-State University Research Excellence,No.SUR/2022/001952.
文摘Recently,we read a mini-review published by Jeyaraman et al.The article explored the optimal methods for isolating mesenchymal stromal cells from adipose tissue-derived stromal vascular fraction(SVF).Key factors include tissue source,processing techniques,cell viability assessment,and the advantages/disadvantages of autologous vs allogeneic use.The authors emphasized the need for standardized protocols for SVF isolation,ethical and regulatory standards for cell-based therapy,and safety to advance mesenchymal stromal cell-based therapies in human patients.This manuscript shares our perspective on SVF isolation in canines.We discussed future directions to potentiate effective regenerative medicine therapeutics in human and veterinary medicine.
文摘BACKGROUND Adipose-derived stem cells(ADSCs)and the stromal vascular fraction(SVF)have garnered substantial interest in regenerative medicine due to their potential to treat a wide range of conditions.Traditional enzymatic methods for isolating these cells face challenges such as high costs,lengthy processing time,and regulatory complexities.AIM This systematic review aimed to assess the efficacy and practicality of nonenzymatic,mechanical methods for isolating SVF and ADSCs,comparing these to conventional enzymatic approaches.METHODS Following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines,a comprehensive literature search was conducted across multiple databases.Studies were selected based on inclusion criteria focused on non-enzymatic isolation methods for SVF and ADSCs from adipose tissue.The risk of bias was assessed,and a qualitative synthesis of findings was performed due to the methodological heterogeneity of the included studies.RESULTS Nineteen studies met the inclusion criteria,highlighting various mechanical techniques such as centrifugation,vortexing,and ultrasonic cavitation.The review identified significant variability in cell yield and viability,and the integrity of isolated cells across different non-enzymatic methods compared to enzymatic procedures.Despite some advantages of mechanical methods,including reduced processing time and avoidance of enzymatic reagents,the evidence suggests a need for optimization to match the cell quality and therapeutic efficacy achievable with enzymatic isolation.CONCLUSION Non-enzymatic,mechanical methods offer a promising alternative to enzymatic isolation of SVF and ADSCs,potentially simplifying the isolation process and reducing regulatory hurdles.However,further research is necessary to standardize these techniques and ensure consistent,high-quality cell yields for clinical applications.The development of efficient,safe,and reproducible non-enzymatic isolation methods could significantly advance the field of regenerative medicine.
文摘Background: Regenerative medicine holds promise for treating degenerative and inflammatory conditions like osteoarthritis (OA). However, the complex molecular mechanisms of OA and the limitations of current therapies remain challenges. Adipose-derived stem cells (ADSC) and stromal vascular fraction (SVF) are gaining attention for OA treatment due to their abundance in adipose tissue. The literature delineates two primary techniques for SVF extraction from adipose tissue: enzymatic digestion and mechanical methods. The Lipocube Hybrid SVF presents a straightforward and secure mechanical isolation method for SVF, enhancing its regenerative potential for various applications. Purpose: This study aims to provide valuable insights into the potential of Lipocube Hybrid SVF as a regenerative therapy for OA, contributing to the broader understanding of its applicability in addressing this debilitating condition. Method: To assess the effectiveness and safety of the Lipocube Hybrid SVF, we have designed a comparative study that evaluates cellular activity and viability, phenotypic characterization, and differentiation potential. The in vitro activity of mechanically isolated SVF is compared to the established gold standard enzymatic digestion method. After in vitro studies, Lipocube Hybrid mechanical isolation method was used to isolate SVF and applied in 42 knee and 7 hip joints of 28 patients with Grade II, Grade III, and Grade IV OA. Results: The Lipocube Hybrid group had slightly lower viable cell numbers but higher cell viability. Flow cytometry analysis showed the Lipocube Hybrid group exhibited more favorable markers for regenerative potential and reduced inflammatory response. Additionally, both groups demonstrated successful osteogenic differentiation, with the Lipocube Hybrid group excelling in chondrogenic and adipogenic differentiation. The clinical application of the Lipocube Hybrid SVF in OA patients resulted in significant improvements in WOMAC and VAS scores across different OA grades. Conclusions: This comparative study was conducted to evaluate the effectiveness and safety of the Lipocube Hybrid SVF, which has shown promise in laboratory settings, for different stages of osteoarthritis. The study findings provide valuable insights into the potential of Lipocube Hybrid SVF as a regenerative therapy for OA, highlighting its suitability for addressing this debilitating condition.
文摘AIM: To investigate the role of vascular endothelial growth factor (VEGF/and its receptors VEGFR-1 and 2 in the growth and differentiation of gastrointestinal strornal tumors (GISTs). METHODS: Thirty-three GISTs, 15 leiomyomas and 6 schwannomas were examined by immunohistochemistry in this study. RESULTS: VEGF protein was expressed in the cytoplasm of tumor cells, and VEGFRol and 2 were expressed both in the cytoplasm and on the membrane of all tumors. Irnrnunohistochernical staining revealed that 26 GISTs (78.8%), 9 leiornyornas (60.0%) and 3 schwannornas (50.0%/were positive for VEGF; 24 GISTs (72.7%/, 12 leiornyornas (80.0%) and 4 schwannornas (66.7%) were positive for VEGFR-1; 30 GISTs (90.9%/, 5 leiornyornas (33.3%/and 4 schwannornas (66.7%) were positive for VEGFR-2. VEGFR-2 expression was statistically different between GISTs and leiomyomas (P 〈 0.0001). However, there was no correlation between the expression of VEGF pathway componenets and the clinical risk categories. CONCLUSION: Our results suggest that the VEGF pathway may play an important role in the differentiation of GISTs, leiomyomas and schwannomas.
文摘AIM: To estimate whether S-TI571 inhibits the expression of vascular endothelial growth factor (VEGF) in the gastrointestinal stromal tumor (GIST) cells. METHODS: We used GIST cell line, GIST-T1. It has a heterogenic 57-bp deletion in exon 11 to produce a mutated c-KIT, which results in constitutive activation of c-KIT. Cells were treated with/without STI571 or stem cell factor (SCF). Transcription and expression of VEGF were determined by RT-PCR and flow cytometry or Western blotting, respectively. Activated c-KIT was estimated by immunoprecipitation analysis. Cell viability was determined by PITT assay. RESULTS: Activation of c-KIT was inhibited by STI571 treatment. VEGF was suppressed at both the transcriptional and translational levels in a temporal and dose-dependent manner by STI571. SCF upregulated the expression of VEGF and it was inhibited by S-13571. STI571 also reduced the cell viability of the GIST-T1 cells, as determined by PTT assay. CONCLUSION: Activation of c-KIT in the GIST-T1 regulated the expression of VEGF and it was inhibited by ST571. STI571 has antitumor effects on the GIST cells with respect to not only the inhibition of cell growth, but also the suppression of VEGF expression.
文摘BACKGROUND The development of fully functional small diameter vascular grafts requires both a properly defined vessel conduit and tissue-specific cellular populations.Mesenchymal stromal cells(MSCs) derived from the Wharton's Jelly(WJ) tissue can be used as a source for obtaining vascular smooth muscle cells(VSMCs),while the human umbilical arteries(h UAs) can serve as a scaffold for blood vessel engineering.AIM To develop VSMCs from WJ-MSCs utilizing umbilical cord blood platelet lysate.METHODS WJ-MSCs were isolated and expanded until passage(P) 4. WJ-MSCs were properly defined according to the criteria of the International Society for Cell and Gene Therapy. Then, these cells were differentiated into VSMCs with the use of platelet lysate from umbilical cord blood in combination with ascorbic acid,followed by evaluation at the gene and protein levels. Specifically, gene expression profile analysis of VSMCs for ACTA2, MYH11, TGLN, MYOCD, SOX9,NANOG homeobox, OCT4 and GAPDH, was performed. In addition,immunofluorescence against ACTA2 and MYH11 in combination with DAPI staining was also performed in VSMCs. HUAs were decellularized and served as scaffolds for possible repopulation by VSMCs. Histological and biochemical analyses were performed in repopulated h UAs.RESULTS WJ-MSCs exhibited fibroblastic morphology, successfully differentiating into"osteocytes", "adipocytes" and "chondrocytes", and were characterized by positive expression(> 90%) of CD90, CD73 and CD105. In addition, WJ-MSCs were successfully differentiated into VSMCs with the proposed differentiation protocol. VSMCs successfully expressed ACTA2, MYH11, MYOCD, TGLN and SOX9. Immunofluorescence results indicated the expression of ACTA2 and MYH11 in VSMCs. In order to determine the functionality of VSMCs, h UAs were isolated and decellularized. Based on histological analysis, decellularized h UAs were free of any cellular or nuclear materials, while their extracellular matrix retained intact. Then, repopulation of decellularized h UAs with VSMCs was performed for 3 wk. Decellularized h UAs were repopulated efficiently by the VSMCs. Biochemical analysis revealed the increase of total hydroyproline and s GAG contents in repopulated h UAs with VSMCs. Specifically, total hydroxyproline and s GAG content after the 1 st, 2 nd and 3 rd wk was 71 ± 10, 74 ± 9 and 86 ± 8 μg hydroxyproline/mg of dry tissue weight and 2 ± 1, 3 ± 1 and 3 ± 1μg s GAG/mg of dry tissue weight, respectively. Statistically significant differences were observed between all study groups(P<0.05).CONCLUSION VSMCs were successfully obtained from WJ-MSCs with the proposed differentiation protocol. Furthermore, h UAs were efficiently repopulated by VSMCs. Differentiated VSMCs from WJ-MSCs could provide an alternative source of cells for vascular tissue engineering.
基金Supported by National Natural Science Foundation of China,No.82070801,No.82100858,No.82073227China Postdoctoral Science Foundation,No.2020M671661+1 种基金Jiangsu Province Science Foundation for Youths,No.BK20200569Jiangsu Province Research Founding for Postdoctoral,No.1412000016.
文摘Osteoarthritis(OA)is considered to be a highly heterogeneous disease with progressive cartilage loss,subchondral bone remodeling,and low-grade inflammation.It is one of the world's leading causes of disability.Most conventional clinical treatments for OA are palliative drugs,which cannot fundamentally cure this disease.The stromal vascular fraction(SVF)from adipose tissues is a heterogeneous cell population.According to previous studies,it contains a large number of mesenchymal stem cells,which have been used to treat OA with good therapeutic results.This safe,simple,and effective therapy is expected to be applied and promoted in the future.In this paper,the detailed pathogenesis,diagnosis,and current clinical treatments for OA are introduced.Then,clinical studies and the therapeutic mechanism of SVF for the treatment of OA are summarized.
文摘AIM:To characterize the implications of vascular endothelial growth factor(VEGF)-A in stromal cells and colorectal cancer and the expression of VEGF-A splice variants.METHODS:VEGF-A expression in tumor and stromal cells from 165 consecutive patients with colorectal cancer was examined by immunohistochemistry.The association between VEGF-A expression status and clinicopathological factors was investigated.Twenty freshfrozen samples were obtained for laser capture microdissection to analyze the splice variants of VEGF-A.RESULTS:VEGF-A was expressed in 53.9% and 42.4% of tumor and stromal cells,respectively.VEGF-A expression in tumor cells(t-VEGF-A) was associated with advanced clinical stage(stage 0,1/9;stage 1,2/16;stage 2,32/55;stage 3,38/66;stage 4,16/19,P < 0.0001).VEGF-A expression in stromal cells(s-VEGF-A) increased in the earlier clinical stage(stage 0,7/9;stage 1,6/16;stage 2,33/55;stage 3,22/66;stage 4,5/19;P = 0.004).Multivariate analyses for risk factors of recurrence showed that only s-VEGF-A expression was an independent risk factor for recurrence(relative risk 0.309,95% confidence interval 0.141-0.676,P = 0.0033).The five-year disease-free survival(DFS) rates of t-VEGF-A-positive and-negative cases were 51.4% and 62.9%,respectively.There was no significant difference in t-VEGF-A expression status.The five-year DFS rates of s-VEGF-A-positive and-negative cases were 73.8% and 39.9%,respectively.s-VEGFA-positive cases had significantly better survival than s-VEGF-A-negative cases(P = 0.0005).Splice variant analysis revealed that t-VEGF-A was mainly composed of VEGF165 and that s-VEGF-A included both VEGF165 and VEGF165b.In cases with no venous invasion(v0),the level of VEGF165b mRNA was significantly higher(v0 204.5 ± 122.7,v1 32.5 ± 36.7,v2 2.1 ± 1.7,P = 0.03).The microvessel density tended to be lower in cases with higher VEGF165b mRNA levels.CONCLUSION:s-VEGF-A appears be a good prognostic factor for colorectal cancer and includes VEGF165 and VEGF165b.
基金the Science and Technology Research Program of Shenzhen,No. 200802005
文摘BACKGROUND:Studies have demonstrated that bone marrow stromal cells (BMSCs) undergo neuronal differentiation under certain in vitro conditions.However,very few inducers of BMSC differentiation have been used in clinical application.The effects of vascular endothelial growth factor (VEGF) on in vitro neuronal differentiation of BMSCs remain poorly understood.OBJECTIVE:To investigate the effect of VEGF on neuronal differentiation of BMSCs in vitro,and to determine the best VEGF concentration for experimental induction.DESIGN,TIME AND SETTING:In vitro comparative study was performed at the Central Laboratory and Laboratory of Male Reproductive Medicine,Shenzhen Hospital of Peking University from October 2008 to August 2009.MATERIALS:Recombinant human VEGF165 was purchased from Peprotech Asia,Rehovot,Israel.Neuron-specific enolase (NSE) was purchased from Beijing Biosynthesis Biotechnology,China.METHODS:BMSCs were harvested from adult Sprague Dawley rats.The passaged cells were pre-induced with 10 ng/mL basic fibroblast growth factor for 24 hours,followed by differentiation induction with 0,5,10,and 20 ng/mL VEGF,respectively.MAIN OUTCOME MEASURES:Morphological changes in BMSCs prior to and following VEGF induction.Expression of NSE following induction was determined by immunocytochemistry.RESULTS:Shrunken,round cells,with a strong refraction and thin bipolar or multipolar primary and secondary branches were observed 3 days after induction with 5,10,and 20 ng/mL VEGF.However,these changes were not observed in the control group.At 10 days after induction,the number of NSE-positive cells was greatest in the 10 ng/mL VEGF-treated group (P〈 0.05).The number of NSE-positive cells was least in the control group at 3 and 10 days post-induction (P〈 0.05).Moreover,the number of NSE-positive cells was greater at 10 days compared with at 3 days after induction (P〈 0.05).CONCLUSION:Of the VEGF concentrations tested,10 ng/mL induced the greatest number of neuronal-like cells in vitro from BMSCs.
文摘The increasing implementation of multicentre studies has led to a need for the optimization of a method that allows for accurate post-hoc analysis of patient biological samples. Assessment of total cell number, viability and immunophenotype can present logistical challenges which can be aided by batch processing. The increased sample storage time that this requires necessitates the use of reagents to preserve cellular integrity, viability and immunophenotype. TransFix is a stabilising reagent that has been developed for the preservation of cell numbers and cell marker expression in peripheral whole blood for up to ten days. This study investigated the use of TransFix reagent for the preservation of the stromal vascular fraction (SVF) of collagenase digested adipose tissue. It was demonstrated that TransFix was suitable for accurately measuring nucleated SVF cell numbers for up to seven days as well as back calculating original cell viability. It also stabilised three CD markers commonly used to identify populations within SVF (CD90, CD31 and CD45) for up to seven days. There was no significant difference between the number of CD90, CD31 and CD45 positive cells after stabilisation at Day 7 compared to Day 0 unstabilised samples. The results suggest that TransFix can be used to preserve a biological mixed cell population from human adipose-derived SVF for up to seven days for accurate post-hoc analysis.
文摘Lipotransfer has become a powerful regenerative tool,largely because of its cellular components,the stromal vascular fraction(SVF).However,the clinical separation of cells with collagenase is strictly legislated.In 2017,Yao et al.postulated a novel fat-derived product mechanically concentrating SVF cells and an extracellular matrix(ECM)and named it stromal vascular fraction gel(SVF-gel).This review discussed the protocol of SVF-gel and its component as well as its inner structure.The histologic examination and the retention rate after the transplantation of SVF-gel were also rendered.Moreover,we summed up the rejuvenating and regenerative use of SVF-gel and introduced its possible mechanism.
文摘Objective To investigate the effect of adipose stromal vascular fraction cells(SVFs)on the survival rate of fat ransplantation.Methods 0.5mL autologous fat tissue was mixed with: ① DiI-labeled autologous SVFs (Group A);②
文摘BACKGROUND Gastrointestinal stromal tumors(GISTs)are considered the most common mesenchymal tumors of the gastrointestinal tract.Microvessel density(MVD)constitutes a direct method of vascularity quantification and has been associated with survival rates in multiple malignancies.AIM To appraise the effect of MVD on the survival of patients with GIST.METHODS This study adhered to Systematic reviews and Meta-Analyses guidelines and the Cochrane Handbook for Systematic Reviews of Interventions.Electronic scholar databases and grey literature repositories were systematically screened.The Fixed Effects or Random Effects models were used according to the Cochran Q test.RESULTS In total,6 eligible studies were identified.The pooled hazard ratio(HR)for disease free survival(DFS)was 8.52(95%CI:1.69-42.84,P=0.009).The odds ratios of disease-free survival between high and low MVD groups at 12 and 60 mo did not reach statistical significance.Significant superiority of the low MVD group in terms of DFS was documented at 36 and 120 mo(OR:8.46,P<0.0001 and OR:22.71,P=0.0003,respectively)as well as at metastases rate(OR:0.11,P=0.0003).CONCLUSION MVD significantly correlates with the HR of DFS and overall survival rates at 36 and 120 mo.Further prospective studies of higher methodological quality are required.