Skip to main content
Log in

Wet milling of large quantities of human excision adipose tissue for the isolation of stromal vascular fraction cells

  • Published:
Cytotechnology Aims and scope Submit manuscript

Abstract

The isolation of stromal vascular fraction (SVF) cells from excised human adipose tissue, for clinical or research purposes, implies the tedious and time consuming process of manual mincing prior to enzymatic digestion. Since no efficient alternative technique to this current standard procedure has been proposed so far, the aim of this study was to test a milling procedure, using two simple, inexpensive and commercially available manual meat grinders, to process large amounts of adipose tissue. The procedure was assessed on adipose tissue resections from seven human donors and compared to manual mincing with scalpels. The processed adipose tissues were digested and the resulting SVF cells compared in terms of number, clonogenicity and differentiation capacity. After 10 min of processing, either device tested yielded on average sixfold more processed material for subsequent cell isolation than manual mincing. The isolation yield of SVF cells (isolated cells per ml of adipose tissue), their viability, phenotype, clonogenicity and osteogenic/adipogenic differentiation capacity, tested by production of mineralized matrix and lipid vacuoles, respectively, were comparable. This new method is practical and inexpensive and represents an efficient alternative to the current standard for large scale adipose tissue resection processing. A device based on the milling principle could be embedded within a streamlined system for isolation and clinical use of SVF cells from adipose tissue excision.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  • Barbero A, Ploegert S, Heberer M, Martin I (2003) Plasticity of clonal populations of dedifferentiated adult human articular chondrocytes. Arthritis Rheumatol 48:1315–1325

    Article  CAS  Google Scholar 

  • Bianco P (2014) Mesenchymal stem cells. Annu Rev Cell Dev Biol 30:677–704

    Article  CAS  Google Scholar 

  • Bianco P, Robey PG (2015) Skeletal stem cells. Development 142:1023–1027

    Article  CAS  Google Scholar 

  • Bishop A, Hong P, Bezuhly M (2014) Autologous fat grafting for the treatment of velopharyngeal insufficiency: state of the art. J Plast Reconstr Aesthet Surg 67:1–8

    Article  CAS  Google Scholar 

  • Borowski DW, Gill TS, Agarwal AK, Tabaqchali MA, Garg DK, Bhaskar P (2015) Adipose tissue-derived regenerative cell-enhanced lipofilling for treatment of cryptoglandular fistulae-in-ano: the ALFA technique. Surg Innov 22:593–600

    Article  Google Scholar 

  • CDC Guideline for disinfection and Sterilization in Healthcare Facilities (2008). https://www.cdc.gov/infectioncontrol/pdf/guidelines/disinfection-guidelines.pdf. Accessed 17 June 2017

  • Choi YS, Dusting GJ, Stubbs S, Arunothayaraj S, Han XL, Collas P, Morrison WA, Dilley RJ (2010) Differentiation of human adipose-derived stem cells into beating cardiomyocytes. J Cell Mol Med 14:878–889

    Article  CAS  Google Scholar 

  • Emnett RJ, Kaul A, Babic A, Geiler V, Regan D, Gross G, Akel S (2016) Evaluation of tissue homogenization to support the generation of GMP-compliant mesenchymal stromal cells from the umbilical cord. Stem Cells Int 2016:3274054

    Article  Google Scholar 

  • Fraser JK, Zhu M, Wulur I, Alfonso Z (2008) Adipose-derived stem cells. Methods Mol Biol 449:59–67

    Google Scholar 

  • Gimble JM, Grayson W, Guilak F, Lopez MJ, Vunjak-Novakovic G (2011) Adipose tissue as a stem cell source for musculoskeletal regeneration. Front Biosci (Sch Ed) 1:69–81

    Article  Google Scholar 

  • Güven S, Mehrkens A, Saxer F, Schaefer DJ, Martinetti R, Martin I, Scherberich A (2011) Engineering of large osteogenic grafts with rapid engraftment capacity using mesenchymal and endothelial progenitors from human adipose tissue. Biomaterials 32:5801–5809

    Article  Google Scholar 

  • Güven S, Karagianni M, Schwalbe M, Schreiner S, Farhadi J, Bula S, Bieback K, Martin I, Scherberich A (2012) Validation of an automated procedure to isolate human adipose tissue-derived cells by using the Sepax® technology. Tissue Eng Part C Methods 18:575–582

    Article  Google Scholar 

  • Hennig T, Lorenz H, Thiel A, Goetzke K, Dickhut A, Geiger F, Richter W (2007) Reduced chondrogenic potential of adipose tissue derived stromal cells correlates with an altered TGFbeta receptor and BMP profile and is overcome by BMP-6. J Cell Physiol 211:682–691

    Article  CAS  Google Scholar 

  • Huang SH, Wu SH, Chang KP, Lin CH, Chang CH, Wu YC, Lee SS, Lin SD, Lai CS (2015) Alleviation of neuropathic scar pain using autologous fat grafting. Ann Plast Surg 74:S99–S104

    Article  CAS  Google Scholar 

  • Klinger M, Caviggioli F, Vinci V, Salval A, Villani F (2010) Treatment of chronic posttraumatic ulcers using autologous fat graft. Plast Reconstr Surg 126:154e–155e

    Article  Google Scholar 

  • Kølle SF, Fischer-Nielsen A, Mathiasen AB, Elberg JJ, Oliveri RS, Glovinski PV, Kastrup J, Kirchhoff M, Rasmussen BS, Talman ML, Thomsen C, Dickmeiss E, Drzewiecki KT (2013) Enrichment of autologous fat grafts with ex vivo expanded adipose tissue-derived stem cells for graft survival: a randomised placebo-controlled trial. Lancet 382:1113–1120

    Article  Google Scholar 

  • Largo RD, Tchang LA, Mele V, Scherberich A, Harder Y, Wettstein R, Schaefer DJ (2014) Efficacy, safety and complications of autologous fat grafting to healthy breast tissue: a systematic review. J Plast Reconstr Aesthet Surg 67:437–448

    Article  Google Scholar 

  • Lin K, Matsubara Y, Masuda Y, Togashi K, Ohno T, Tamura T, Toyoshima Y, Sugimachi K, Toyoda M, Marc H, Douglas A (2008) Characterization of adipose tissue-derived cells isolated with the celution system. Cytotherapy 10:417–426

    Article  CAS  Google Scholar 

  • Mehrkens A, Saxer F, Güven S, Hoffmann W, Müller AM, Jakob M, Weber FE, Martin I, Scherberich A (2012) Intraoperative engineering of osteogenic grafts combining freshly harvested, human adipose-derived cells and physiological doses of bone morphogenetic protein-2. Eur Cell Mater 28:308–319

    Article  Google Scholar 

  • Meijer GJ, de Bruijn JD, Koole R, van Blitterswijk CA (2008) Cell based bone tissue engineering in jaw defects. Biomaterials 29:3053–3061

    Article  CAS  Google Scholar 

  • Mitchell JB, McIntosh K, Zvonic S, Garrett S, Floyd ZE, Kloster A, Di Halvorsen Y, Storms RW, Goh B, Kilroy G, Wu X, Gimble JM (2006) Immunophenotype of human adipose-derived cells: temporal changes in stromal-associated and stem cell-associated markers. Stem Cells 24:376–378

    Article  Google Scholar 

  • Müller AM, Mehrkens A, Schäfer DJ, Jaquiery C, Güven S, Lehmicke M, Martinetti R, Farhadi I, Jakob M, Scherberich A, Martin I (2010) Towards an intraoperative engineering of osteogenic and vasculogenic grafts from the stromal vascular fraction of human adipose tissue. Eur Cell Mater 3:127–135

    Article  Google Scholar 

  • Navarro A, Marín S, Riol N, Carbonell-Uberos F, Miñana MD (2014) Human adipose tissue-resident monocytes exhibit an endothelial-like phenotype and display angiogenic properties. Stem Cell Res Ther 14:50

    Article  Google Scholar 

  • Nishimura S, Manabe I, Nagasaki M, Hosoya Y, Yamashita H, Fujita H, Ohsugi M, Tobe K, Kadowaki T, Nagai R, Sugiura S (2007) Adipogenesis in obesity requires close interplay between differentiating adipocytes, stromal cells, and blood vessels. Diabetes 56:1517–1526

    Article  CAS  Google Scholar 

  • Ong WK, Sugii S (2013) Adipose-derived stem cells: fatty potentials for therapy. Int J Biochem Cell Biol 45:1083–1086

    Article  CAS  Google Scholar 

  • Osinga R, Menzi NR, Tchang LA, Caviezel D, Kalbermatten DF, Martin I, Schaefer DJ, Scherberich A, Largo RD (2015) Effects of intersyringe processing on adipose tissue and its cellular components: implications in autologous fat grafting. Plast Reconstr Surg 135:1618–1628

    Article  CAS  Google Scholar 

  • Osinga R, Di Maggio N, Todorov A, Allafi N, Barbero A, Laurent F, Schaefer DJ, Martin I, Scherberich A (2016) Generation of a bone organ by human adipose-derived stromal cells through endochondral ossification. Stem Cells Transl Med 5:1090–1097

    Article  CAS  Google Scholar 

  • Pallua N, Baroncini A, Alharbi Z, Stromps JP (2014) Improvement of facial scar appearance and microcirculation by autologous lipofilling. J Plast Reconstr Aesthet Surg 67:1033–1037

    Article  CAS  Google Scholar 

  • Pasquale P, Gaetano M, Giovanni DO, Luigi C, Gilberto S (2015) Autologous fat grafting in facial volumetric restoration. J Craniofac Surg 26:756–759

    Article  Google Scholar 

  • Salem HK, Thiemermann C (2010) Mesenchymal stromal cells: current understanding and clinical status. Stem Cells 31:585–596

    Google Scholar 

  • Saxer F, Scherberich A, Todorov A, Studer P, Miot S, Schreiner S, Güven S, Tchang LA, Haug M, Heberer M, Schaefer DJ, Rikli D, Martin I, Jakob M (2016) Implantation of stromal vascular fraction progenitors at bone fracture sites: from a rat model to a first-in-man study. Stem Cells 34:2956–2966. https://doi.org/10.1002/stem.2478

    Article  CAS  Google Scholar 

  • Scherberich A, Müller AM, Schäfer DJ, Banfi A, Martin I (2010) Adipose tissue-derived progenitors for engineering osteogenic and vasculogenic grafts. J Cell Physiol 225:348–353

    Article  CAS  Google Scholar 

  • Scherberich A, Di Maggio ND, McNagny KM (2013) A familiar stranger: CD34 expression and putative functions in SVF cells of adipose tissue. World J Stem Cells 26:1–8

    Article  Google Scholar 

  • Tamura E, Fukuda H, Tabata Y, Nishimura M (2008) Use of the buccal fat [corrected] pad for vocal cord augmentation. Acta Otolaryngol 128:219–224

    Article  Google Scholar 

  • Volat F, Bouloumié A (2013) Steroid hormones and the stroma-vascular cells of the adipose tissue. Horm Mol Biol Clin Investig 15:5–10

    CAS  Google Scholar 

Download references

Funding

This work was supported by the Swiss National Science Foundation (SNF Grant No. 310030-138519, to A.S. and I.M.).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Ivan Martin.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This study was approved by the local ethical committee (Ethikkommission beider Basel [EKBB], Ref. 78/07 extended in 2009). All seven donors gave written informed consent.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (MP4 9888 kb)

Supplementary material 2 (MP4 14188 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Menzi, N., Osinga, R., Todorov, A. et al. Wet milling of large quantities of human excision adipose tissue for the isolation of stromal vascular fraction cells. Cytotechnology 70, 807–817 (2018). https://doi.org/10.1007/s10616-018-0190-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10616-018-0190-z

Keywords

Navigation