Comparison of fetal bovine serum and platelet-rich plasma on human lipoaspirate-derived mesenchymal stem cell proliferation.

Background: Lipoaspirate-derived stem cells (LSCs) are very promising for regenerative medicine, e.g. to treat acute myocard infarction. Fetal bovine serum (FBS) is commonly used to propagate the LSCs. However, for its clinical application, FBS contains xeno-proteins that are potential to elicit immune rejection in patients. Platelet rich plasma (PRP) is one of the candidates to replace FBS. This study was aimed to compare the proliferation of LSCs cultured in 5% PRP, 10% PRP, and FBS containing medium (MesenCult®). Methods: LSCs were cultured in 5% PRP/DMEM, 10% PRP/DMEM, and MesenCult®. After the primary culture reached its confluency, cells were harvested using TrypLE Select and seeded (around 20,000 viable cells) in new vessels in the same media. Passages were done until passage-5, with six replications. Population doubling time (PDT) of the three groups were analyzed using Kruskal Wallis test. Results: LSCs showed different proliferation rates when cultured in 5% PRP/DMEM, 10% PRP/DMEM, and MesenCult®. PDT of the three experimental groups in passage 1-5 were significanly different (p < 0.05), with the lowest rank was cultured in medium of 10% PRP/DMEM. Conclusion: The results suggest that 10% PRP/DMEM can be used as an alternative to replace FBS in LSC culture.

Application of a modified method for stem cell isolation from lipoaspirates in a basic lab.

Aim Lipoaspirate, a wasted by product from liposuction procedure recently has been shown to contain abundant mesenchymal stem cells (MSCs). MSCs have been studied in many research areas to regenerate many cell lineages including, myogenic, cardiomyogenic, and angiogenic lineages. The large quantity of MSCs in lipoaspirate, makes it an attractive source for stem cells used in research and clinical applications. A simplified method which is suitable to be performed in a basic laboratory will facilitate development of stem cell research in developing countries. Therefore the outcomes from this study are expected to encourage the progress of stem cell research in Indonesia. Methods Lipoaspirate was digested using collagenase type I, followed by a basic filtration method. Purification of MSCs was done by cell culture for 2-3 days followed by supernatant removal. To confirm the homogenous population of MSCs, an analysis using flowcytometry was performed based on the MSCs minimal criteria developed by Mesenchymal and Tissue Stem Cell Committee of the International Society of Cell Therapy. Resuts MSCs were able to be obtained at 16.41 ± 8.22 x 108 cells per 120 ml lipoaspirate. The cultured cells showed fibroblastic morphology which is characteristic for MSCs and were able to be purified from non-MSCs cells. This was confirmed by flowcytometry assay showing expression of CD105 and the absence of HLA-Class II, CD 45, CD 34, CD14, and CD19. Conclusions This study has shown that it was feasible to isolate messenchymal stem cell from human lipoaspirate. The procedure was practicable to be performed within a basic laboratory.

Effect of Lipoaspirate Cell Autograft on Proliferation and Collagen Synthesis of Diabetic Fibroblasts in Vitro

PURPOSE: Human lipoaspirate cells are relatively easy to obtain in large quantities without cell culture. The aim of this in vitro pilot study is to identify the effects of cell therapy using uncultured lipoaspirate cells on cell proliferation and collagen synthesis of diabetic fibroblasts, which are the major contributing factors in wound healing. METHODS: In order to get diabetic fibroblasts, dermis tissues were obtained from foot skin of diabetic patients who underwent debridements or toe amputations (n = 4). In order to isolate lipoaspirate cells, the same diabetic patients' abdominal adipose tissues were obtained by liposuction. The diabetic fibroblasts were co-cultured with or without autogenous lipoaspirate cells using porous culture plate insert. Initial numbers of the lipoaspirate cells and diabetic fibroblasts seeded were 15,000 cells/well, respectively. For cell proliferation assay, two treatment groups were included. In group I, diabetic fibroblasts were cultured with the insert having no cells, which serves as a control. In group II, the lipoaspirate cells were added in the culture plate insert. For collagen synthesis assay, one additional group (group III) was included for a reference, in which diabetic fibroblasts were not seeded in the well and only lipoaspirate cells inside the insert were incubated without diabetic fibroblasts. RESULTS: One hundred to one hundred sixty thousand lipoaspirate cells were isolated per ml of aspirated adipose tissue. After 3-day incubation, the mean cell numbers in group I and II were 17,294/well and 22,163/well. The mean collagen level in group I, II, and III were 29, 41, and 2 ng/mL, respectively. These results imply that both cell proliferation and collagen synthesis in the lipoaspirate cell treatment group were 28 and 44 percents higher than in the control group, respectively (p < 0.05). CONCLUSION: Uncultured lipoaspirate cell autografts may stimulate the wound healing activity of diabetic fibroblasts.