Plasma Rich In Growth Factors Eye Drops Research Articles

Beneficial Effects of Plasma Rich in Growth Factors (PRGF) Versus Autologous Serum and Topical Insulin in Ocular Surface Cells

Purpose In the last few decades, several blood derived products such as platelet-rich plasma (PRP), plasma rich in growth factors (PRGF) and autologous serum (AS) have been used for the treatment of ocular surface disorders. Recently, insulin has been proposed to be used as an alternative for the treatment of ocular surface diseases. The aim of this study was to evaluate the biological potential of PRGF eye drops in comparison with AS and insulin on ocular surface cells. Methods Blood from three healthy young donors was collected to obtain autologous serum (AS) eye drops and plasma rich in growth factors (PRGF) eye drops. Insulin (Actrapid®) was diluted at 1 and 0.2 IU/mL. The biological potential of PRGF, AS and insulin was assessed by proliferation in HCE, HK and HConF cells. Wound healing assay was performed in HCE cells after incubation with the different treatments. HConF and HK cells were differentiated to myofibroblast after treatment with 2.5 ng/mL of TGF-β1 and then incubated with all treatments. Results PRGF eye drops induced significantly higher proliferation rate compared to AS or insulin in HConF and HK cells, but not in HCE cells. In addition, the percentage of wound healing area was significantly reduced after PRGF treatment in comparison with AS or insulin treatment. Furthermore, PRGF significantly reduced the number of myodifferentiated cells compared to AS and insulin at both concentrations analyzed. Conclusion The results obtained in the present study show that PRGF increases the biological activity of the ocular surface cells and reduces the expression of fibrosis marker compared to insulin or AS. Translational relevance The present study suggests that plasma rich in growth factors eye drops are a more effective therapy than insulin and autologous serum eye drops for the treatment of ocular surface diseases.

A multicenter report of the use of plasma rich in growth factors (PRGF) for the treatment of patients with ocular surface diseases in North America

PurposeTo investigate the efficacy and safety of plasma rich in growth factors (PRGF) eyedrops in the management of patients with ocular surface diseases in North America. MethodsMulticenter interventional case series of patients using PRGF eyedrops for the first time. A cohort of patients was analyzed for corneal staining score at initial visit and at 3 months of therapy with PRGF. Another cohort responded to a 10-item questionnaire that evaluated patients' satisfaction and safety, which included the symptom assessment questionnaire in dry eye (SANDE) score, after 6 months of PRGF treatment. ResultsA total of 153 patients were analyzed. Of these, 102 were reviewed for corneal epitheliopathy and 99 patients responded to the questionnaire. The mean (±SD) age of the population was 63.7 ± 17 years and 72.5% were female. The clinical indications for PRGF usage were dry eye (60%), neurotrophic keratopathy (15%), dormant corneal ulcers (12%), limbal stem cell deficiency (10%), and cicatrizing conjunctivitis (4%). At the final visit, 74.3% of patients showed an improvement of their corneal staining. Those who had punctate epithelial erosions or epithelial defects were reduced from 76.5% to 47% and 23.5% to 7.8% respectively (p < 0.0001). Symptoms, measured via SANDE score, significantly decreased from a median of 90 to 34.6 out of 100 points on follow-up (p < 0.0001). Only one patient (0.98%) complained of ocular burning sensation as a side effect. ConclusionsThis multicentric study demonstrates the safety and efficacy of the use of PRGF for treating signs and symptoms in patients with significant ocular surface diseases.

VEGF-A in Serum and Plasma Rich in Growth Factors (PRGF) Eye Drops.

Vascular endothelial growth factor (VEGF)-A, the most abundant subtype of the VEGF family in the eye, plays an important role in corneal homeostasis due to its ability to mediate corneal nerve repair. Repeated intravitreal anti-VEGF injections were shown to significantly reduce corneal nerve density, which might negatively affect corneal homeostasis and lead to a neuropathic dry eye disease. Currently, there are two effective modalities to treat dry eye while supplying VEGF to the ocular surface: serum eye drops (SED) and eye drops manufactured from plasma rich in growth factors (PRGF). The purpose of this study was to measure the VEGF-A concentration in SED and PRGF eye drops. Ten healthy volunteers donated blood on two separate occasions, 2 - 8 days apart. Thus, a total of 20 blood samples were processed to obtain both SED and PRGF. Concentrations of VEGF-A were quantified by a Simple Plex platform run in triplicate. The VEGF-A concentration in SED and PRGF was very similar between the two blood samples drawn from one individual donor but showed substantial interindividual variability. However, in all 20 samples, VEGF concentrations were substantially higher in SED samples (mean 238.7 ± 146.6 pg/mL) compared to PRGF samples (mean 67.4 ± 46.3 pg/mL). Based upon the analysis of variance (ANOVA) model for the measured concentrations with fixed effects for specimen (SED vs. PRGF) and subject, the mean difference between the SED and PRGF concentration was 168.1 pg/mL (95% confidence interval: [142.4, 193.9], p < 0.001). Our study showed that the VEGF concentration was higher in SED than in PRGF. This is an important finding, particularly for potential treatment of dry eye disease in patients with neuropathic eye disease, especially in patients that received repeated anti-VEGF intravitreal injections, or in patients with Sjögren's disease, where the level of VEGF in tears might be pathologically decreased. Hypothetically, VEGF might be needed to restore ocular surface homeostasis. Although growing evidence has shown that VEGF-A plays an important role in corneal homeostasis, only a randomized prospective clinical trial will show whether supplying VEGF-A to the ocular surface might successfully restore the corneal homeostasis and overcome the problem of corneal neuropathy in these patients. For such a trial, based on our results, an undiluted SED should be preferred over a PRGF due to the higher content of VEGF-A.

Proteomic Characterization of Plasma Rich in Growth Factors and Undiluted Autologous Serum.

Over the last three decades, there has been special interest in developing drugs that mimic the characteristics of natural tears for use it in the treatment of several ocular surface disorders. Interestingly, the composition of blood plasma is very similar to tears. Therefore, different blood-derived products like autologous serum (AS) and plasma rich in growth factors (PRGF) have been developed for the treatment of diverse ocular pathologies. However, scarce studies have been carried out to analyze the differences between both types of blood-derived products. In the present study, blood from three healthy donors was drawn and processed to obtain AS and PRGF eye drops. Then, human corneal stromal keratocytes (HK) were treated with PRGF or undiluted AS. Proteomic analysis was carried out to analyze and characterize the differential protein profiles between PRGF and AS, and the differentially expressed proteins in HK cells after PRGF and AS treatment. The results obtained in the present study show that undiluted AS induces the activation of different pathways related to an inflammatory, angiogenic, oxidative stress and scarring response in HK cells regarding PRGF. These results suggest that PRGF could be a better alternative than AS for the treatment of ocular surface disorders.

Short- and Long-Term Stability of Plasma Rich in Growth Factors Eye Drops.

To analyze whether plasma rich in growth factors (PRGFs) eye drops preserve their activity and biological properties after storage for 9 and 12 months at -20°C, and at 4°C, and at room temperature (RT) for 3 and 7 days in comparison to fresh samples (t0). PRGF eye drops were obtained from 6 healthy donors. Then, they were stored for 9 and 12 months at -20°C. At each time, different PRGF eye drops samples were thawed and maintained at RT or at 4°C for 3 and 7 days. Platelet-derived growth factor-AB, epidermal growth factor, transforming growth factor-β1, vascular endothelial growth factor, angiopoietin-1, and thrombospondin-1 were analyzed at each time and temperature of storage. In addition, the pH level, the microbial contamination, and the proliferative potential on primary human corneal stromal fibroblasts human keratocytes of each obtained PRGF eye drops were also evaluated. All growth factor levels were preserved at each time and storage condition. No differences were observed on the human keratocytes proliferation after treatment with PRGF eye drops at any studied time or temperature. No microbial contamination was observed in any of the PRGF eye drops. Finally, the pH levels increased significantly after 9 and 12 months of storage at -20°C compared with t0. PRGF eye drops can be stored for up to 12 months without reduction of the main growth factors and proteins and without any microbial contamination. Furthermore, the biological activity of the PRGF eye drops is maintained after storing for 3 and 7 days at 4°C or at RT.

Development and Optimization of Freeze-Dried Eye Drops Derived From Plasma Rich in Growth Factors Technology.

PurposeTo investigate whether plasma rich in growth factors (PRGF) eye drops maintain their biological potential after a freeze drying process. The addition of a lyoprotectant like trehalose was also evaluated.MethodsBlood from three healthy donors was collected to obtain eye drops by PRGF technology. The resultant eye drops were divided in four groups: PRGF, freeze-dried PRGF (PRGF lyo), and PRGF lyophilized mixed with 2,5% trehalose (PRGF lyo+2.5T) or 5% trehalose (PRGF lyof+5T). Chemical and biological characteristics were evaluated. Photorefractive keratectomy was performed on C57BL/6 mice which were divided in three treatment groups: control, PRGF, and PRGF lyo. Corneal wound healing and haze formation were evaluated macroscopically. Eyes were collected at 1, 2, 3, and 7 days after surgery, and were processed for histologic studies.ResultsThe pH values of PRGF samples increased significantly after the lyophilization process. Osmolarity levels increased significantly in PRGF samples mixed with trehalose in comparison with PRGF samples without protectants. The freeze drying process maintained growth factors levels as well as the biological properties of PRGF eye drops even without the use of lyoprotectants. PRGF lyo treatment significantly decreased the re-epithelialization time and haze formation in photorefractive keratectomy-treated corneas regarding PRGF and control groups. Furthermore, the PRGF lyo group significantly decreased the number of smooth muscle actin-positive cells in comparison with the control group at each time of the study and at days 2 and 3 in the PRGF group.ConclusionsThe freeze drying process preserves the protein and growth factor content as well as the biological properties of PRGF eye drops, even without the use of protectants. Freeze-dried PRGF eye drops accelerate corneal tissue regeneration after photorefractive keratectomy in comparison with the control group.Translational RelevanceOur study shows the feasibility to preserve the biological capability of PRGF eye drops as freeze-dried formulation, avoiding the addition of protectants.

Differential profile of protein expression on human keratocytes treated with autologous serum and plasma rich in growth factors (PRGF).

PurposeThe main objective of this study is to compare the protein expression of human keratocytes treated with Plasma rich in growth factors (PRGF) or autologous serum (AS) and previously induced to myofibroblast by TGF-β1 treatment.MethodsBlood from healthy donor was collected and processed to obtain AS and PRGF eye drops. Blood derivates were aliquoted and stored at -80°C until use. Keratocyte cells were pretreated for 60 minutes with 2.5 ng/ml TGF-β1. After that, cells were treated with PRGF, AS or with TGF-β1 (control). To characterize the proteins deregulated after PRGF and AS treatment, a proteomic approach that combines 1D-SDS–PAGE approach followed by LC–MS/MS was carried out.ResultsResults show a catalogue of key proteins in close contact with a myofibroblastic differentiated phenotype in AS treated-cells, whereas PRGF-treated cells show attenuation on this phenotype. The number of proteins downregulated after PRGF treatment or upregulated in AS-treated cells suggest a close relationship between AS-treated cells and cytoskeletal functions. On the other hand, proteins upregulated after PRGF-treatment or downregulated in AS-treated cells reveal a greater association with processes such as protein synthesis, proliferation and cellular motility.ConclusionThis proteomic analysis helps to understand the molecular events underlying AS and PRGF-driven tissue regeneration processes, providing new evidence that comes along with the modulation of TGF-β1 activity and the reversion of the myofibroblastic phenotype by PRGF.

Plasma rich in growth factors eye drops to treat secondary ocular surface disorders in patients with glaucoma.

PurposeTo evaluate the efficacy and safety of plasma rich in growth factors (PRGF) eye drops in patients with glaucoma with secondary ocular surface disorders (OSDs) due to surgeries and topical hypotensive drugs use.Materials and methodsA retrospective case-series study design was used including six patients (eight eyes) diagnosed with glaucoma who received surgical (nonpenetrating deep sclerectomy and/or trabeculectomy) and medical treatments (hypotensive eye drops) to control intraocular pressure (IOP) and who developed secondary OSDs, unresponsive to conventional treatments. Patients were treated with PRGF eye drops (four times a day). Outcome measures were ocular surface disease index (OSDI), best-corrected visual acuity (BCVA, in logarithm of the minimum angle of resolution), visual analog scale (VAS), frequency and severity of symptoms, and IOP. The safety of the treatment was also evaluated.ResultsSix patients (seven eyes with open-angle glaucoma and one eye with uveitic glaucoma) treated with PRGF eye drops were evaluated. Mean age was 71 years (SD=7.2, range 58–79 years). Five were female and one was male. The mean treatment time was 21.8 weeks (SD=9.0, range 12–36 weeks). The mean time to reach closure of the corneal ulcer was 14.5 (SD=5.5) weeks. A statistical significant reduction in OSDI scale (50.6%), VAS frequency (53.1%), VAS severity (42.0%), and a 41.8% improvement in BCVA were observed (p<0.05). IOP also decreased by 16.6% (p=0.010). Only one of the six patients reported itching in both eyes as an adverse event (AE); however, the patient continued with the PRGF eye drops until the end of therapy; the remaining patients did not report any AEs during the follow-up period.ConclusionsIn patients with glaucoma and secondary OSDs refractive to conventional treatments, the treatment with PRGF eye drops could be considered a possible therapeutic option, because it demonstrates an improvement in the signs and symptoms of the ocular surface, as well as a better control of the IOP. This is an initial research work that can open doors for future research to confirm these findings.

Plasma Rich in Growth Factors for the Treatment of Dry Eye after LASIK Surgery

Purpose: The aim of this study was to evaluate the use of plasma rich in growth factors (PRGF) eye drops in patients with dry eye disease after laser-assisted in situ keratomileusis (LASIK) surgery. Material and Methods: This is a longitudinal, retrospective, comparative, and descriptive study of 77 eyes of 42 patients with dry eye disease following LASIK surgery. This study was designed to evaluate the efficacy of PRGF treatment compared to conventional therapy (control group). Outcome measures including signs and symptoms of dry eye disease were evaluated before and after treatment. The percentage of change before and after treatment for each clinical variable measured was compared between both groups. Results: There were 1–4 treatment cycles with PRGF eye drops (1 cycle = 6 weeks). Results showed a statistically significant improvement in the Ocular Surface Disease Index (38.12%), visual analogue scale scores for frequency (41.89%) and severity (42.47%), and the Schirmer test scores (88.98%) after PRGF treatment (p < 0.05). No adverse events were reported after PRGF treatment. Conclusions: These results suggest that PRGF eye drops are effective for the improvement of dry eye symptoms in patients who underwent LASIK surgery in comparison to the conventional therapy. The treatment with PRGF is an alternative for patients who suffer from postoperative dry eye.

Treatment of patients with neurotrophic keratitis stages 2 and 3 with plasma rich in growth factors (PRGF-Endoret) eye-drops.

To provide preliminary data about efficacy and safety of plasma rich in growth factors (PRGF) eye-drops in neurotrophic keratitis (NK) and to analyze the possible influence of certain variables on treatment outcomes. This retrospective study included patients with stages 2-3 of NK treated with PRGF eye-drops. Primary endpoint was the resolution time of corneal ulcer defect. Outcome measures including percentage of ulcer closure at 4weeks, Ocular Surface Disease Index (OSDI), Best-Corrected Visual Acuity (BCVA) and Visual Analogue Scale (VAS) were also evaluated before and after treatment with PRGF. The influence of some patients' clinical variables on results was assessed. Safety assessment was also performed reporting all adverse events. Thirty-eight treated eyes in a total of thirty-one patients were evaluated, of which five cases had no prior response to autologous serum treatment. Most cases (97.4%) achieved the complete resolution of corneal defect/ulcer. Mean resolution time was 11.4weeks (SD=13.7). A statistical significant (p<0.05) reduction in OSDI (60.9%), VAS frequency (59.9%), VAS severity (57.0%) and improvement in BCVA (52.8%) was observed. The results were stratified according to the pathology stage and to the identified potential effect modifiers variables. Only one adverse event was reported in one patient (2.6%). PRGF eye-drops could be a safe and effective therapeutic option for patients with stages 2-3 of NK, showing high rates of corneal defect/ulcer resolution in short times, either in reducing signs and symptoms of NK, and therefore preventing the progression of NK to greater ocular complications.

PRGF exerts more potent proliferative and anti-inflammatory effects than autologous serum on a cell culture inflammatory model

Ocular graft versus host disease (oGVHD) is part of a systemic inflammatory disease that usually affects ocular surface tissues manifesting as a dry eye syndrome. Current treatments provide unsatisfactory results. Blood-derived products, like plasma rich in growth factors (PRGF) emerge as a potential therapy for this disease. The purpose of this study was to evaluate the tissue regeneration and anti-inflammatory capability of PRGF, an autologous platelet enriched plasma eye-drop, compared to autologous serum (AS) obtained from oGVHD patients on ocular surface cells cultured in a pro-inflammatory environment. PRGF and AS were obtained from four GVHD patients. Cell proliferation and inflammation markers, intercellular adhesion molecule-1 (ICAM-1) and cyclooxygenase-2 (COX-2), were measured in corneal and conjunctival fibroblastic cells cultured under pro-inflammatory conditions and after treatment with PRGF or AS eye drops. Moreover, cell proliferation increased after treatment with PRGF and AS, though this enhancement in the case of keratocytes was significantly higher with PRGF. PRGF eye drops showed a significant reduction of both inflammatory markers with respect to the initial inflammatory situation and to the AS treatment. Our results concluded that PRGF exerts more potent regenerative and anti-inflammatory effects than autologous serum on ocular surface fibroblasts treated with pro-inflammatory IL-1β and TNFα.

Safety and Efficacy of Autologous Plasma Rich in Growth Factors Eye Drops for the Treatment of Evaporative Dry Eye

Purpose: To provide preliminary information about the safety and efficacy of plasma rich in growth factors (PRGF) in the treatment of evaporative dry eye (EDE) and analyze the influence of certain variables on treatment outcomes. Methods: This retrospective study included patients with EDE treated between January 2011 and December 2013. Outcome measures including signs and symptoms of the disease were evaluated before and after treatment. The influence of some of the patient's clinical variables on results was assessed. Safety assessment was also performed. Results: Eighty-three patients with a total of 156 eyes were evaluated. Statistically significant reductions in the Ocular Surface Disease Index (38.2%), best-corrected visual acuity (27.4%), Visual Analogue Scale scores for frequency (32%) and severity (34%), and a significant improvement in the Schirmer test scores (177.5%) were observed (p < 0.05). The results were stratified according to the identified potential effect modifiers. Only four adverse events were reported. All of them were mild and resolved within a few days. Conclusions: Results suggest that PRGF eye drops could be a safe and effective treatment in reducing EDE signs and symptoms. When treating patients, the possible influence of some clinical variables must be taken into account.

Autologous Plasma Rich in Growth Factors Eyedrops in Refractory Cases of Ocular Surface Disorders

Purpose: Preliminary information about the safety and efficacy of plasma rich in growth factors (PRGF) eyedrops in the treatment of refractory cases of diverse ocular surface disorders (OSDs) is presented here. Material and Methods: This retrospective cohort study included cases with OSDs refractory to previous treatment with conventional treatments or autologous serum or cyclosporine, and treated with PRGF eyedrops. The signs and symptoms of ocular surface disorders [using the ocular surface disease index (OSDI), best-corrected visual acuity (BCVA), visual analog scale (VAS) frequency and VAS severity] were evaluated before and after treatment with PRGF. A safety assessment was also performed reporting all adverse events or complications. Results: Forty-one patients with a total of 80 treated eyes were evaluated. Statistically significant reductions in the OSDI scale (39.27%), VAS frequency (38.9%) and VAS severity (40.3%), and a significant improvement in BCVA (54.86%) were all observed (p < 0.05). The results were stratified according to the identified potential effect modifiers. There were only two adverse events (eye redness and eyelid inflammation), which were reported as mild and resolved in a few days. Conclusions: PRGF eyedrops could be a safe and effective treatment option for refractory cases of OSDs. When treating patients the possible influence on the results of some clinical variables must be taken into account.

Preservation of Biological Activity of Plasma and Platelet-Derived Eye Drops After Their Different Time and Temperature Conditions of Storage.

To analyze whether plasma rich in growth factors (PRGF) eye drops preserve their biological characteristics and activity after storage for 3 and 6 months at -20°C, at 4°C, and at room temperature for 72 hours, compared with fresh samples (t0). Blood from 6 healthy donors was harvested and centrifuged to obtain PRGF free of leukocytes. Resulting PRGF eye drops were stored for 3 and 6 months at -20°C. At each time, 2 aliquots were maintained at room temperature or at 4°C for 72 hours. Platelet-derived growth factor-AB, transforming growth factor-β1, vascular endothelial growth factor, epidermal growth factor, insulin-like growth factor-1, angiopoietin-1, and thrombospondin-1 were quantified at each time and temperature of storage. Also, the effect of PRGF eye drops on proliferation of primary human keratocytes was evaluated. All the analyzed growth factor levels remained constant at each time and storage condition. No differences were observed in the proliferative activity of keratocytes after treatment with PRGF eye drops at any studied time or temperature. Finally, there was no microbial contamination in any of the PRGF eye drops. The preservation of the PRGF eye drops at -20°C for up to 3 and 6 months does not mean reduction of the main growth factors and proteins implicated in ocular surface wound healing. Eye drop characteristics and in vitro biological activity were not affected by their usage and conservation for 72 hours at 4°C or at room temperature.

Plasma rich in growth factors (PRGF) eye drops stimulates scarless regeneration compared to autologous serum in the ocular surface stromal fibroblasts

Autologous serum (AS) eye drops was the first blood-derived product used for the treatment of corneal pathologies but nowadays PRGF arises as a novel interesting alternative to this type of diseases. The purpose of this study was to evaluate and compare the biological outcomes of autologous serum eye drops or Plasma rich in growth factors (PRGF) eye drops on corneal stromal keratocytes (HK) and conjunctival fibroblasts (HConF). To address this, blood from healthy donors was collected and processed to obtain autologous serum (AS) eye drops and plasma rich in growth factors (PRGF) eye drops. Blood-derivates were aliquoted and stored at −80°C until use. PDGF-AB, VEGF, EGF, FGFb and TGF-β1 were quantified. The potential of PRGF and AS in promoting wound healing was evaluated by means of proliferation and migration assays in HK and HConF. Fibroblast cells were induced to myofibroblast differentiation after treatment with 2.5ng/mL of TGF-β1. The capability of PRGF and AS to prevent and inhibit TGF-β1-induced differentiation was evaluated. Results showed significant higher levels of all growth factors analyzed in PRGF eye drops compared to AS. Moreover, PRGF eye drops enhanced significantly the biological outcomes of both HK and HConF, and reduced TGF-β1-induced myofibroblast differentiation in contrast to autologous serum eye drops (AS). In summary, these results suggest that PRGF exerts enhanced biological outcomes than AS. PRGF may improve the treatment of ocular surface wound healing minimizing the scar formation compared to AS. Results obtained herein suggest that PRGF protects and reverses the myofibroblast phenotype while promotes cell proliferation and migration.

Effects of heat-treatment on plasma rich in growth factors-derived autologous eye drop

We have developed and characterized a new type of plasma rich in growth factors (PRGF) derived eye-drop therapy for patients suffering from autoimmune diseases. To determine the concentration of several growth factors, proteins, immunoglobulins and complement activity of the heat-inactivated eye-drop and to study its biological effects on cell proliferation and migration of different ocular surface cells, blood from healthy donors was collected, centrifuged and PRGF was prepared avoiding the buffy coat. The half volume of the obtained plasma supernatant from each donor was heat-inactivated at 56 °C for 1 h (heat-inactivated PRGF). The concentration of several proteins involved on corneal wound healing, immunoglubolins G, M and E and functional integrity of the complement system assayed by CH50 test were determined. The proliferative and migratory potential of inactivated and non-inactivated PRGF eye drops were assayed on corneal epithelial cells (HCE), keratocytes (HK) and conjunctival fibroblasts (HConF). Heat-inactivated PRGF preserves the content of most of the proteins and morphogens involved in its wound healing effects while reduces drastically the content of IgE and complement activity. Heat-inactivated PRGF eye drops increased proliferation and migration potential of ocular surface cells with regard to PRGF showing significant differences on proliferation and migration rate of HCE and HConF respectively. In summary, heat-inactivation of PRGF eye drops completely reduced complement activity and deceased significantly the presence of IgE, maintaining the biological activity of PRGF on ocular surface cells.