title: Co-culture of mesenchymal-like stromal cells derived from human foreskin permits long term propagation and differentiation of human embryonic stem cells.
abstract: Among the different parameters governing the successful derivation and expansion of human embryonic stem cells (hESC), feeder layers play the most important role. Human feeders in form of human mesenchymal stromal cells (hMSCs) and human foreskin fibroblasts (HFFs) lay the foundation for eradication of animal-derived hESC culture system. In this study we explored the potential of human foreskin derived mesenchymal like stromal cells (HF-MSCs) to support self renewal and pluripotency of hESC. The MSCs isolated from human foreskin were found to be resistant to standard concentrations and duration of mitomycin-C treatment. Growth pattern, gene profiling (Oct-4, Nanog, Sox-2, Rex-1), cytoskeletal protein expression (vimentin, nestin) and tri-lineage differentiation potential into adipocytes, chondrocytes and osteocytes confirmed their mesenchymal stromal cell status. Further, the HF-MSCs were positive for CD105, CD166, CD73, CD44, CD90, SSEA-4, and negative for CD34, CD45, HLA-DR cell-surface markers and were found to exhibit BM-MSC-like characteristics. hESC lines co-cultured with HF-MSC feeders showed expression of expected pluripotent transcription factors Oct-4, Nanog, Sox-2, GDF-3, Rex-1, STELLAR, ABCG2, Dppa5, hTERT; surface markers SSEA-4, TRA-1-81 and maintained their cytogenetic stability during long term passaging. These novel feeders also improved the formation of embryoid bodies (EBs) from hESC which produced cell types representing three germ layers. This culture system has the potential to aid the development of clinical-grade hESCs for regenerative medicine and drug screening. Further, we envisage foreskin can serve as a valuable source of alternative MSCs for specific therapeutic applications.
authors.affiliation: Stempeutics Research Malaysia Sdn. Bhd., Technology Park Malaysia, 57000 Kuala Lumpur, Malaysia.
authors.forename: Murali Krishna
authors.initials: MK
authors.lastname: Mamidi
authors.forename: Rajarshi
authors.initials: R
authors.lastname: Pal
authors.forename: Nor Azah Binti
authors.initials: NA
authors.lastname: Mori
authors.forename: Greetha
authors.initials: G
authors.lastname: Arumugam
authors.forename: Saratha Thevi
authors.initials: ST
authors.lastname: Thrichelvam
authors.forename: Puteri J
authors.initials: PJ
authors.lastname: Noor
authors.forename: Hj Mohamad Farouk
authors.initials: HM
authors.lastname: Abdullah
authors.forename: Pawan Kumar
authors.initials: PK
authors.lastname: Gupta
authors.forename: Anjan Kumar
authors.initials: AK
authors.lastname: Das
authors.forename: Zubaidah
authors.initials: Z
authors.lastname: Zakaria
authors.forename: Ramesh
authors.initials: R
authors.lastname: Bhonde
mesh_terms: D050156:Adipogenesis / Q000235:genetics; D000293:Adolescent; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D002648:Child; D018920:Coculture Techniques; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D052816:Foreskin / Q000166:cytology; D006801:Humans; D008297:Male; D008648:Mesoderm / Q000166:cytology* / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism; D017154:Stromal Cells / Q000166:cytology / Q000378:metabolism; D055815:Young Adult