Results from this study demonstrate that the double centrifugation tube method is a reliable technique for producing equine P-PRP, such as it was demonstrated previously . This method was initially evaluated in a mixed group of 26 horses of different breeds, ages and, gender . In that study PLT concentration was lower and WBC concentration was higher in P-PRP  in comparison with the hematological results from our study. However, it was no possible to find significant differences associated with the intrinsic variables  such as was found in the present research. Thus, results from our study (using a larger sample size of horses) suggest that centrifugation protocols for producing equine P-PRP and perhaps L-PRP, should be adapted in function of the breed, since possibly the size and weight of PLT and WBC could be different for each equine specific breed.
Some studies suggest that, cellular content will influence the GF concentration in a determined PC . This fact was evidenced when TGF-β1 and PDGF-BB concentrations were measured in the different blood components of our study. However, although there are several equine studies evaluating the GF concentration in PRP, there is conflicting results in the TGF-β1 and PDGF-BB concentrations from equine PG obtained by either manual methods [8, 13] or semi-automated devices [9, 10, 14, 15]. The technique used for GF determination and non-standardized technical aspects inherent to each research, prevent comparing our findings with some of these studies.
However, our GF results could be compared with the TGF-β1 and PDGF-BB concentrations reported by Textor et al. for equine PRP produced by a manual method and a semi-automated device. They reported the concentration for these GF in PRP+NID and compared their release from PRP activated with two collagen type I (COL1) concentrations, 10 and 20 μg/mL . In our study, lower TGF-β1 and PDGF-BB concentrations (mean ± d.s., 3192.7 ± 1395.3 and 1259.7 ± 418.6 pg/mL, respectively), were obtained from P-PRP+NID in comparison with equine PRP+NID (22677 ± 12125 and 4332 ± 2212 pg/mL, respectively) reported by Textor et al.. The difference observed for GF concentrations in these blood components could be associated with a higher number of PLT and WBC concentrated in PRP+NID from Textor et al.’s study . On the other hand, PRP activated with COL1 produced a very lower GF release in comparison with the TGF-β1 and PDGF-BB concentration released from P-PRG activated with CG at 6 h. The findings of our study suggest that CG produces a complete release of GF from P-PRG in the first 6 h post-activation. In contrast, COL1 produced a very weak GF release from PRP .
Our study demonstrated that intrinsic factors such as, breed, gender and age influence the cellular composition of P-PRP and GF content in P-PRG. These findings are novel and should be considered in the clinical use of these bioproducts. However, it is important to consider that these observations should also be confirmed for either equine L-PRP or L-PRG obtained with semi-automated devices using an adequate simple size of horses to avoid unpowered studies with erroneous conclusions .
The most intriguing result of our study was the significantly higher PDGF concentrations in P-PRP+NID and P-PRG derived from CCH in comparison with ACH. This same aspect was also observed in females in comparison with males and in horses younger than 5 years old in comparison with older horses. To note, CCH is a pony-like breed and our findings could suggest that PDGF-BB from platelets could be a pivotal factor associated with accelerated limb wound healing in ponies in comparison with horses . It is important to bear in mind that anabolic GF, such as insulin like growth factor type I is also highly expressed and produced in young horses in comparison with older ones .The main limitation of our study was associated with the fact that only CCH females were used for gender comparison. However, this is the first time that significant differences between genders for PLT count in P-PRP and PDGF-BB concentrations in P-PRG have been described in horses. At least, two aspects could be useful to explain these differences. First, the present study used a sample size greater than in other equine studies [3, 8–10, 13]. This situation added statistical power to the study and increased the possibility of finding significant differences for the evaluated variables . Second, gender differences have been reported for PLT and WBC counts in human beings with greater concentration of these cells in females . While it is known that hormonal influences could be associated with these hematological gender differences in human beings , we did not measure the concentration of sexual hormones in horses from our study to support this hypothesis.
The correlation analysis from our study suggests that PDGF-BB is one of the main GF contained in equine PLT. On the other hand, TGF-β1concentrations were associated with both PLT and WBC counts. Of note, PDGF-BB concentrations were correlated with the type of breed, gender and the age. These findings suggest that this GF should be used as an indicator of PLT enrichment in PRP and as indicator of equine PRG quality.
Taken together, our data suggests that intrinsic factors, such as breed, sex and age can influence the cellular and GF profile of equine P-PRP/P-PRG. Specific protocols for concentrating platelets and GF by using manual or semi-automated devices should be standardized in function of the equine intrinsic factors. Additional studies are necessary to know if these intrinsic factors could influence the therapeutic potential of P-PRP/P-PRG in horses.