scholarly journals Use of microfiltered vs only disaggregated mesenchymal stem cells from adipose tissue in regenerative medicine

2020 ◽  
Vol 51 (3) ◽  
pp. 152-157
Author(s):  
Fabiano Svolacchia ◽  
Lorenzo Svolacchia
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Bioactive Peptides ◽  
Clinical Use ◽  
Adult Mesenchymal Stem Cells ◽  
Epidermal Regeneration ◽  
Inflammatory Component ◽  
Number Of Cells

Background: Clinical use of adult mesenchymal stem cells (MSCa) in medicine and regenerative surgery is constantly evolving. Adipose tissue-derived stem cells (ADSc) are capable of inducing the production of new extracellular matrix (ECM), deposition of new collagen and early revascularisation. Methods: Flow cytometry was performed for 2 mL of cell colonies harvested from adipose tissue (AT). Comparation has been made of at disaggregated only and the same at disaggregated and microfiltered at 50 mm, 100 mm and 200 mm. Signs of inflammation after dermo-epidermal regeneration session through the mesotherapy method were observed and compared. Results: Even after filtration, significant number of ADSc was collected. An increase in the size of the filter did not always translate into an increase in the number of cells that were found in the microfiltrate. In the non-filtered at disaggregated in both cases, highest number of cells was found, as expected, but at the expense of more pronounced inflammation. Sampling with the 16 Gauge needle produces superior results compared to the cannula in all cases. Conclusion: With this method in medicine and regenerative surgery it will be easier to exploit the growth factors, mRNA, MicroRNA, lipids and bioactive peptides emitted in the MSCa signalling micro-vesicles as they are isolated from the inflammatory component.

OPTIMIZATION of the Cultivation of Mesenchymal STEM CELLS for CLINICAL USE

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4394-4394
Author(s):  
Vanessa de Souza Valim ◽  
Fernanda Oliveira ◽  
Maria Aparecida Lima da Silva ◽  
Bruna Amorin ◽  
Lauro Moraes ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Conflicts Of Interest ◽  
Calf Serum ◽  
Optimal Number ◽  
Third Party ◽  
Clinical Use ◽  
Clinical Grade ◽  
Plating Density ◽  
Number Of Cells

Abstract Abstract 4394 Introduction and objectives: Studies with mesenchymal stem cells (MSCs) have shown its benefits in hematology, mainly for Graft-versus-host disease (Lancet 371:1579–86, 2008), with three unsettled matters: (1)MSCs expansion in medium supplemented with Fetal Calf Serum (FCS) and its risk of xenoreaction (Blood 89:776–9, 1997); (2)The optimal number of cells needed for therapy is not yet defined, but there is an empirical indication for 2×106células/Kg with the need to optimize expansion, number and time wise; and (3)the utilization of third party donors. This study was designed to determine the superiority or no-inferiority of the Platelet Lysates (PL) over FCS on the expansion of MSC, the optimal cell plating density and days between each pass, and to investigate if in our conditions total nucleated cells (TNC) obtained from the washouts of HSCT explants can expand to be used at clinical grade. Methods and Results: TNC were removed from the filters and bags used in the HSCT (Cytotherapy 11:403–13, 2009) and after the first passage were plated in different concentrations (2000/cm2, 3000/cm2, 4000/cm2, 5000/cm2, 6000/cm2 and 7000/cm2) with 10% FBS or 10% PL, and the number of days reach 80% of confluence was observe (Transfusion, 49:2680–5, 2009). Next, cultures with the same plating density were fed either with 10% PL or 10% FCS and were trypsinized after seven days and counted to analyze how much they have grown in that time period. And finally, cultures were allowed to growth up to Passage 3 (P3) to test the ability to obtain clinical grade number of cells. The proliferation of mesenchymal stem cells in the presence of PL and SFB was averaged 11.88 and 2.5 times, respectively, in a period of 7 days (p = 0.005). The highest concentration of plating cells using PL, took less time (6 days) to reach confluence as compared with the three lower (7.55 to 8.55 days) (p = 0.005), and at P3 with PL we obtained from 10×109 up to 10 × 1011 cells. Conclusion: This study suggests that the PL is the best choice as a supplement to expand MSC, and allow the proliferation of a sufficient number of MSC at P3 for clinical use obtained from the washouts of HSCT explants. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Comparative Analysis of Apoptotic Resistance of Mesenchymal Stem Cells Isolated from Human Bone Marrow and Adipose Tissue

2012 ◽  
Vol 2012 ◽  
pp. 1-8 ◽  
Author(s):  
Gökhan Ertaş ◽  
Ertan Ural ◽  
Dilek Ural ◽  
Ayça Aksoy ◽  
Güliz Kozdağ ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Stem Cells ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Serum Deprivation ◽  
Human Bone ◽  
Induced Apoptosis

Aim. Mesenchymal stem cells (MSCs) isolated from human bone marrow (hBM) and adipose tissue (hAT) are perceived as attractive sources of stem cells for cell therapy. The aim of this study was to compare MSCs from hBM and hAT for their immunocytochemistry staining and resistance to in vitro apoptosis. Methods. In our study, we investigated the antiapoptotic ability of these MSCs toward oxidative stress induced by hydrogen peroxide (H2O2) and serum deprivation. Results were assessed by MTT and flow cytometry. All experiments were repeated a minimum of three times. Results. Flow cytometry and MTT analysis revealed that hAT-MSCs exhibited a higher resistance toward H2O2-induced apoptosis (n=3, hBM-hAT viability H2O2  58.43±1.24–73.02±1.44, P<0.02) and to serum-deprivation-induced apoptosis at days 1 and 4 than the hBM-MSCs (n=3, hAT-hBM absorbance, resp., day 1: 0.305±0.027–0.234±0.015, P=0.029, day 4: 0.355±0.003–0.318±0.007, P=0.001, and day 7: 0.400±0.017–0.356±0.008, P=0.672). hAT-MSCs showed superior tolerance to oxidative stress triggered by 2 mmol/L H2O2 and also have superior antiapoptosis capacity toward serum-free culture. Conclusion. In this study we found that hAT-MSCs are more resistant to in vitro apoptosis.

scholarly journals Characterization of mesenchymal stem cells derived from equine adipose tissue

2013 ◽  
Vol 65 (4) ◽  
pp. 939-945 ◽  
Author(s):  
A.M. Carvalho ◽  
A.L.M. Yamada ◽  
M.A. Golim ◽  
L.E.C. Álvarez ◽  
L.L. Jorge ◽  
...  
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Mhc Class Ii ◽  
Class Ii ◽  
Specific Staining ◽  
Alizarin Red ◽  
Lineage Differentiation ◽  
Mixed Breed

Stem cell therapy has shown promising results in tendinitis and osteoarthritis in equine medicine. The purpose of this work was to characterize the adipose-derived mesenchymal stem cells (AdMSCs) in horses through (1) the assessment of the capacity of progenitor cells to perform adipogenic, osteogenic and chondrogenic differentiation; and (2) flow cytometry analysis using the stemness related markers: CD44, CD90, CD105 and MHC Class II. Five mixed-breed horses, aged 2-4 years-old were used to collect adipose tissue from the base of the tail. After isolation and culture of AdMSCs, immunophenotypic characterization was performed through flow cytometry. There was a high expression of CD44, CD90 and CD105, and no expression of MHC Class II markers. The tri-lineage differentiation was confirmed by specific staining: adipogenic (Oil Red O), osteogenic (Alizarin Red), and chondrogenic (Alcian Blue). The equine AdMSCs are a promising type of adult progenitor cell for tissue engineering in veterinary medicine.

scholarly journals Expression of CD34 marker in the adipose tissue derived stem cells

2018 ◽  
Vol 5 (1) ◽  
Author(s):  
Phuc Van Pham ◽  
Ngoc Bich Vu ◽  
Van Hong Tran
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Hematopoietic Stem Cells ◽  
Adipose Derived Stem Cells ◽  
Differentiation Potential ◽  
Hematopoietic Stem ◽  
Cd34 Expression

Introduction: Adipose-derived stem cells (ADSCs) are considered as mesenchymal stem cells (MSCs). Indeed, they display all characteristics of MSCs that compliant with the minimal criteria of MSCs suggested by Domonici et al. (2006). However, some recent studies showed that ADSCs contain the subpopulation that was positive with CD34 marker – a marker of hematopoietic stem cells. This study aimed to analyze and determine the expression of CD34 marker in ten samples of ADSCs obtained from 10 donors. Methods: All ADSC samples were isolated and expanded according to the published previous protocols. They were confirmed as the MSCs with some markers and differentiation potential, excepting the CD34 expression. Then they were cultured and analyzed the expression of CD34 by flow cytometry at passage 3, 5, 7 and 9. Results: The results showed that expression of CD34 in ADSCs was different between donors and their passages that accounted from 1.21% to 23.38%. Conclusion: These results suggested that ADSCs are not ‘truly” MSCs like MSCs from bone marrow.

scholarly journals Improvement of an Effective Protocol for Directed Differentiation of Human Adipose Tissue-Derived Adult Mesenchymal Stem Cells to Corneal Endothelial Cells

2021 ◽  
Vol 22 (21) ◽  
pp. 11982
Author(s):  
Cadenas-Martin Marta ◽  
Moratilla Adrian ◽  
Fernández-Delgado Jorge ◽  
Arnalich-Montiel Francisco ◽  
Maria P. De Miguel
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Corneal Transplantation ◽  
Human Adipose Tissue ◽  
Directed Differentiation ◽  
Human Donor ◽  
Adult Mesenchymal Stem Cells ◽  
Human Adscs ◽  
Induced Pluripotent

Corneal disease affects 12.5 million individuals worldwide, with 2 million new cases each year. The standard treatment consists of a corneal transplantation from a human donor; however, the worldwide demand significantly exceeds the available supply. Lamellar endothelial keratoplasty, the replacement of only the endothelial layer of the cornea, can partially solve the problem. Progressive efforts have succeeded in expanding hCECs; however, the ability to expand hCECs is still limited, and new sources of CECs are being sought. Crucial advances have been achieved by the directed differentiation of embryonic or induced pluripotent stem cells, but these cells have disadvantages, such as the use of oncogenes, and are still difficult to establish. We aimed to transfer such knowledge to obtain hCECs from adipose tissue-derived adult mesenchymal stem cells (ADSC) by modifying four previously published procedures. We present several protocols capable of the directed differentiation of human ADSCs to hCECs. In our hands, the protocol by Ali et al. was the best adapted to such differentiation in terms of efficiency, time, and financial cost; however, the protocol by Wagoner et al. was the best for CEC marker expression. Our results broaden the type of cells of autologous extraocular origin that could be employed in the clinical setting for corneal endothelial deficiency.

scholarly journals Adipose-derived stem cells in the treatment of knee osteoarthritis: from extraction methods to the preparation of the transplant

2022 ◽  
Vol 11 (1) ◽  
pp. e24111124699
Author(s):  
Laynna de Carvalho Schweich-Adami ◽  
Larissa Corrêa Hermeto ◽  
Silvana Marques Caramalac ◽  
Andréia Conceição Milan Brochado Antoniolli-Silva ◽  
Rodrigo Juliano Oliveira
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Knee Osteoarthritis ◽  
Cellular Therapy ◽  
Extraction Methods ◽  
International Federation ◽  
Adipose Derived Stem Cells ◽  
Real Presence ◽  
Number Of Cells

Introduction: Recent studies have investigated the use of adipose tissue as source of mesenchymal stem cells in the treatment of knee osteoarthritis in humans. However, there are still several protocols being performed. Objective: Analyze the protocols published in the literature in the last ten years and to investigate how they are being carried out and if they are following the criteria adopted by the International Federation for Adipose Therapeutics and Science (IFATS) and the International Society for Cellular Therapy (ISCT). Methodology: Articles from the PubMed, ScienceDirect and Lilacs database published in January / 2010 until the present time, which were evaluated in order to investigate the use of adipose-derived stem cells in the treatment of knee osteoarthritis. Results: Thirty four articles were evaluated in its entiraty. The abdominal area was the most choosen to do the liposuction, however the quantities of adipose tissue removed and the number of cells transplanted was variable.  It is hightlited the enzimatic digestion of adipose tissue with collagenase as extraction method. Only 14 articles complied all the 3 criteria required to prove the real presence of mesenchymal stem cells in the samples that was transplanted. However, all the articles showed improvement of function and pain. Final considerations: Thus, even the results found are promising, the evidence is still limited in humans and the variability of the methodology makes it difficult to standardize the technique, also its implementation as a reference in the treatment of knee osteoarthritis.

scholarly journals Comparative analysis of mouse bone marrow and adipose tissue mesenchymal stem cells for critical limb ischemia cell therapy

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Pegah Nammian ◽  
Seyedeh-Leili Asadi-Yousefabad ◽  
Sajad Daneshi ◽  
Mohammad Hasan Sheikhha ◽  
Seyed Mohammad Bagher Tabei ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Cell Therapy ◽  
Femoral Artery ◽  
Critical Limb Ischemia ◽  
Limb Ischemia

Abstract Introduction Critical limb ischemia (CLI) is the most advanced form of peripheral arterial disease (PAD) characterized by ischemic rest pain and non-healing ulcers. Currently, the standard therapy for CLI is the surgical reconstruction and endovascular therapy or limb amputation for patients with no treatment options. Neovasculogenesis induced by mesenchymal stem cells (MSCs) therapy is a promising approach to improve CLI. Owing to their angiogenic and immunomodulatory potential, MSCs are perfect candidates for the treatment of CLI. The purpose of this study was to determine and compare the in vitro and in vivo effects of allogeneic bone marrow mesenchymal stem cells (BM-MSCs) and adipose tissue mesenchymal stem cells (AT-MSCs) on CLI treatment. Methods For the first step, BM-MSCs and AT-MSCs were isolated and characterized for the characteristic MSC phenotypes. Then, femoral artery ligation and total excision of the femoral artery were performed on C57BL/6 mice to create a CLI model. The cells were evaluated for their in vitro and in vivo biological characteristics for CLI cell therapy. In order to determine these characteristics, the following tests were performed: morphology, flow cytometry, differentiation to osteocyte and adipocyte, wound healing assay, and behavioral tests including Tarlov, Ischemia, Modified ischemia, Function and the grade of limb necrosis scores, donor cell survival assay, and histological analysis. Results Our cellular and functional tests indicated that during 28 days after cell transplantation, BM-MSCs had a great effect on endothelial cell migration, muscle restructure, functional improvements, and neovascularization in ischemic tissues compared with AT-MSCs and control groups. Conclusions Allogeneic BM-MSC transplantation resulted in a more effective recovery from critical limb ischemia compared to AT-MSCs transplantation. In fact, BM-MSC transplantation could be considered as a promising therapy for diseases with insufficient angiogenesis including hindlimb ischemia.

scholarly journals Extracellular Vesicles Do Not Mediate the Anti-Inflammatory Actions of Mouse-Derived Adipose Tissue Mesenchymal Stem Cells Secretome

2021 ◽  
Vol 22 (3) ◽  
pp. 1375
Author(s):  
María Carmen Carceller ◽  
María Isabel Guillén ◽  
María Luisa Gil ◽  
María José Alcaraz
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Extracellular Vesicles ◽  
Nuclear Factor Κb ◽  
Peritoneal Macrophages ◽  
Toll Like Receptor 4 ◽  
Anti Inflammatory ◽  
Phagocytic Response

Adipose tissue represents an abundant source of mesenchymal stem cells (MSC) for therapeutic purposes. Previous studies have demonstrated the anti-inflammatory potential of adipose tissue-derived MSC (ASC). Extracellular vesicles (EV) present in the conditioned medium (CM) have been shown to mediate the cytoprotective effects of human ASC secretome. Nevertheless, the role of EV in the anti-inflammatory effects of mouse-derived ASC is not known. The current study has investigated the influence of mouse-derived ASC CM and its fractions on the response of mouse-derived peritoneal macrophages against lipopolysaccharide (LPS). CM and its soluble fraction reduced the release of pro-inflammatory cytokines, adenosine triphosphate and nitric oxide in stimulated cells. They also enhanced the migration of neutrophils or monocytes, in the absence or presence of LPS, respectively, which is likely related to the presence of chemokines, and reduced the phagocytic response. The anti-inflammatory effect of CM may be dependent on the regulation of toll-like receptor 4 expression and nuclear factor-κB activation. Our results demonstrate the anti-inflammatory effects of mouse-derived ASC secretome in mouse-derived peritoneal macrophages stimulated with LPS and show that they are not mediated by EV.

scholarly journals Derivation, characterization, and in vitro cell regeneration of canine white adipose tissue-derived mesenchymal stem cells obtained from a mesenteric region

2021 ◽  
Vol 82 (1) ◽  
Author(s):  
Anirban Mandal ◽  
Ajeet Kumar Jha ◽  
Dew Biswas ◽  
Shyamal Kanti Guha
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
White Adipose Tissue ◽  
Stromal Vascular Fraction ◽  
Cell Regeneration ◽  
Wound Healing Assay ◽  
Chain Reaction ◽  
Polymerase Chain

Abstract Background The study was conducted to assess the characterization, differentiation, and in vitro cell regeneration potential of canine mesenteric white adipose tissue-derived mesenchymal stem cells (AD-MSCs). The tissue was harvested through surgical incision and digested with collagenase to obtain a stromal vascular fraction. Mesenchymal stem cells isolated from the stromal vascular fraction were characterized through flow cytometry and reverse transcription-polymerase chain reaction. Assessment of cell viability, in vitro cell regeneration, and cell senescence were carried out through MTT assay, wound healing assay, and β-galactosidase assay, respectively. To ascertain the trilineage differentiation potential, MSCs were stained with alizarin red for osteocytes, alcian blue for chondrocytes, and oil o red for adipocytes. In addition, differentiated cells were characterized through a reverse transcription-polymerase chain reaction. Results We observed the elongated, spindle-shaped, and fibroblast-like appearance of cells after 72 h of initial culture. Flow cytometry results showed positive expression for CD44, CD90, and negative expression for CD45 surface markers. Population doubling time was found 18–24 h for up to the fourth passage and 30±0.5 h for the fifth passage. A wound-healing assay was used to determine cell migration rate which was found 136.9 ± 4.7 μm/h. We observed long-term in vitro cell proliferation resulted in MSC senescence. Furthermore, we also found that the isolated cells were capable of differentiating into osteogenic, chondrogenic, and adipogenic lineages. Conclusions Mesenteric white adipose tissue was found to be a potential source for isolation, characterization, and differentiation of MSCs. This study might be helpful for resolving the problems regarding the paucity of information concerning the basic biology of stem cells. The large-scale use of AD-MSCs might be a remedial measure in regenerative medicine.

Sign in / Sign up

Export Citation Format

Share Document