The Effect of PRF on Serum Starved Human Dermal Fibroblast

Abstract

Healing failure on chronic ulcers was suspected due to the decrease of Growth Factors (GFs) supply caused by either GFs trapped in the fibrin, or degraded by protease, or decreased level due to reduction of GFs gene expression. Administration of various GFs can stimulate healing of chronic ulcers. High level of GFs is available in biologic material called Platelet Rich Fibrin (PRF). This study was conducted to have in vitro evidence of PRF effect on GFs-serum starved human dermal fibroblasts as representative cells of chronic ulcers. Human dermal fibroblasts (HDFs) were isolated from foreskin of six boys aged 11-14 years-old. After 24 hours of serum deprivation, HDFs were treated by 100, 50, and 25% PRF lysate diluted in cultured medium. Cellular migration was measured using scratch assay, while cellular viability was measured using MTT assay and collagen deposition was measured using Sirius Red assay. The HDFs of serum starvation group showed significant impairment activities in terms of cellular migration (25%), cellular proliferation (20%), and collagen deposition (10%) (p<0.05). Administration with various levels of PRF lysate could significantly recover those activities (p < 0.05). Because cellular activities of serum starved HDFs is similar with fibroblasts isolated from the bottom of chronic ulcers and administration of various levels of PRF lysate was capable to recover those activities, it can be concluded that PRF is a good biologic material to stimulate healing of chronic ulcers. However, in order to get better evidence based medicine, both pre clinical and clinical studies must be performed. ABSTRAKKegagalan sembuh pada ulkus khronis disebabkan oleh rendahnya suplai growth factor (GF) akibat GF terjebak dalam fibrin, atau dirusak oleh protease bakteri, atau akibat menurunnya ekspresi gena penyandi GF. Pemberian GF dapat memacu penyembuhan ulkus khronis. Berbagai GF ternyata terdapat dalam material biologis seperti fibrin kaya platelet (FKP). Penelitian ini dilakukan untuk memperoleh bukti in vitro bahwa FKP dapat mengembalikan fungsi fibroblast tanpa serum sebagai model fibroblas dari ulkus khronis. Fibroblas dermis manusia (FDM) diisolasi dari kulit kulup 6 anak laki-laki berumur 11-14 tahun. Setelah dibebaskan dari serum selama 24 jam, FDM kemudian diberi 100, 50, 25% lisat FKP dan medium pelarut sebagai kontrol. Selanjutnya, aktivitas selular berupa timbunan kolagen, proliferasi dan migrasi sel diukur berturut-turut dengan metode Sirius Merah, MTT, dan goresan. FDM tanpa serum ternyata secara mengalami penurunan kemampuan menimbun kolagen sebesar 10%, proliferasi sebesar 20% dan penurunan kemampuan migrasi sebesar 25% secara signifikan (p<0,05). Penambahan lisat FKP 111Radiono, The effect of platelet rich fibrin (PRF) on serum starved human dermal fibroblast ternyata dapat memulihkan aktivitas selular FDM tersebut. Dari eksperimen ini dapat disimpulkan bahwa FKP merupakan material biologis yang dapat dikembangkan untuk memacu penyembuhan ulkus khronis. Namun demikian, untuk memperoleh bukt...

Effects of platelet-rich plasma and platelet-poor plasma on human dermal fibroblasts

“…Thereafter, the PRP was diluted with media to concentrations of 100%, 50%, and 25% of PRP and subsequently applied to the challenged fibroblasts in which all samples ameliorated suppressed proliferation. The highest migration capacity was restored via the 50% PRP, and the 100% PRP (no additional medium) restored collagen deposition in challenged fibroblasts [55]. These results propose PRP as an adjunct therapy for chronic ulcers and that higher platelet concentrations proved the most efficacious.…”

Leucocyte-Rich Platelet-Rich Plasma Enhances Fibroblast and Extracellular Matrix Activity: Implications in Wound Healing

“…The aim of this study's three experiments was to explore the effect of leucocytes contained in LR-PRP on a key cell that plays a central role in wound healing namely the fibroblast. In the proliferation experiment, the aim was to determine the effects of LR-PRP on fibroblast which were challenged in serum-free media and consequently ceased proliferation to conserve energy [21,22,67]. This assay identified the optimal fibroblast RNA extraction time to be 12 h; however the time limit of 48 h was insufficient to produce fibroblast proliferation data.…”

“…In order to simulate a fibroblast in the edges of a chronic wound the fibroblasts were deprived of nutrients by being placed in serum-free media. Under these conditions, fibroblasts conserve energy and cease proliferation [ 20 , 21 , 22 ]. The challenged fibroblasts were supplemented with LR-PRP and LP-PPP, and the effects were evaluated.…”

The combination of ADSCs and 10% PRP increases Rb protein expression on senescent human dermal fibroblasts

“…The cells were grown in a solution containing 20% FBS and Dulbecco's Modified Eagle Medium (FBS-DMEM, ThermoFisher). The method used was a modified version of the method used by Takashima (1998), while the senescent HDF method used was that of Radiono et al (2016). 20,21 The HDFs were shifted into a senescent state by replacing the medium with 10% PRP in DMEM (ThermoFisher).…”