The Use of Human Platelet Lysate as a Coating Substance for Adipose-Derived Stem Cell Expansion.

BACKGROUND: Large-scale production of mesenchymal stromal cells is essential for sufficient therapeutic doses in regenerative medicine. However, long-term cultivation encounters limited cell growth and cellular aging. Therefore, an alternative cell culture approach that promotes proliferation and attenuates cell senescence is required. Human platelet lysate (HPL) is a potent supplement for in vitro cell expansion. Applying HPL as a coating material can potentially improve mesenchymal stromal cell cultures. METHOD: To examine the capacity of HPL, it was used to pre-coat a tissue culture plate for in vitro adipose-derived mesenchymal stromal cell expansion. Alterations in biological features of adipose-derived stem cells (ADSCs) were investigated, including cell adhesion assays, cell proliferation, population doubling time, and cellular senescence. RESULTS: ADSCs cultured on HPL-coated plates significantly increased cell adhesion rate, shortened population doubling time, and stimulated cell growth. The senescent cells were significantly decreased in ADSCs cultured in an HPL-coated plate, and the expression levels of senescence-associated genes, including p16, p21, and p53, were downregulated. Furthermore, Western blotting analysis revealed that HPL was enriched with fibronectin and vitronectin, essential cell adhesive proteins. CONCLUSIONS: HPL was effectively used as a coating material for ADSC expansions. Cellular cultivation on the HPL coating is an alternative approach for producing mesenchymal stromal cells.

Application of human platelet lysate in chondrocyte expansion promotes chondrogenic phenotype and slows senescence progression via BMP-TAK1-p38 pathway.

Osteoarthritis (OA) is one of the most common musculoskeletal degenerative. OA treatments are aiming to slow down disease progression; however, lack of cartilage regeneration efficacy. Autologous chondrocyte implantation (ACI) is a promising cartilage-regeneration strategy that uses human articular chondrocytes (HACs) as cellular materials. However, the unreadiness of HACs from prolonged expansion, cellular senescence, and chondrogenic dedifferentiation occurred during conventional expansion, thus, minimizing the clinical efficacy of ACI. We aimed to examine the effects of a human platelet lysate (HPL) as an alternative human-derived HAC medium supplement to overcome the limitations of conventional expansion, and to explain the mechanism underlying the effects of HPL. During passages 2-4 (P2-P4), HPL significantly increased HAC proliferation capacities and upregulated chondrogenic markers. Simultaneously, HPL significantly reduced HAC senescence compared with conventional condition. HACs treated with LDN193189 exhibited a reduction in proliferation capacity and chondrogenic marker expression, whereas the HAC senescence increased slightly. These findings indicated involvement of BMP-2 signaling transduction in the growth-assistive, anti-senescent, and chondrogenic-inductive properties of HPL, which demonstrated its beneficial effects for application as HAC medium supplement to overcome current expansion limitations. Finally, our findings support the roles of platelets in platelet-rich plasma as a promising treatment for patients with OA.

Indoxyl sulfate impairs in vitro erythropoiesis by triggering apoptosis and senescence.

Anemia is a major complication in over 50% of chronic kidney disease (CKD) patients. One of the main causes of anemia in CKD is the reduction of erythropoietin (EPO) synthesis from renal tubular cells. Therefore, first-line treatment of CKD is EPO administration; however, EPO unresponsiveness in several patients is frequently found. More undefined causes of anemia in CKD are under interest, especially uremic toxins, which are a group of solutes accumulated in CKD patients. The highly detectable protein-bound uremic toxin, indoxyl sulfate (IS) was investigated for its effects on in vitro erythropoiesis in this study. CD34+ hematopoietic stem cells were isolated from human umbilical cord blood and differentiated toward erythrocyte lineage for 14 days in various concentrations of IS (12.5, 25, 50, and 100 µg/mL). The effects of IS on cell proliferation, differentiation, apoptosis, and senescence were determined. Cell proliferation was investigated by manual cell counting. Cell surface marker expression was analyzed by flow cytometry. Wright's staining was performed to evaluate cell differentiation capacity. Apoptosis and senescence marker expression was measured using reverse transcription polymerase chain reaction (RT-PCR). TUNEL assay was performed to detect apoptotic DNA fragmentation. Our results demonstrated that IS reduced cell proliferation and impaired erythrocyte differentiation capacity. In addition, this study confirmed the effects of IS on cell apoptosis and senescence during erythropoietic differentiation. Therefore, the promotion of apoptosis and senescence might be one of the possible mechanisms caused by uremic toxin accumulation leading to anemia in CKD patients.