The effects of stromal vascular fraction and platelet rich plasma on bone healing of rat model with chronic renal disease

Present studies show that %10 of the world population have problems related to kidney disease at any level. It is known that chronic kidney disease (CKD) impairs osteoblast/ osteoclast balance and damages bone structure with mechanisms such as diminished mineralization. In this study; we aim to investigate the effects of adipose derived stromal vascular fraction (SVF) and platelet-rich plasma (PRP) on bone healing model in rats with chronic renal failure. In this study; 62 male Sprague- Dawley rats were used and seperated into four groups as, Group I (n:12), Group II (n:12), Group III (n:12), Group IV (n:16). 10 rats were used for harvesting SVF and PRP. All groups except Group I, which was used as healthy control group, had CKD surgery using 5/6 nephrectomy operation. Both groups had intramedullary pin fixation after receiving bone fracture on their left femur with the help of drilling tools. Group II rats were used as control group for CKD. Group III rats received PRP treatment on fracture site. Group IV rats received PRP and SVF treatment on fracture site. Rats were diagnosed and followed as CKD with weight loss parameters and hemoglobin, urea and creatinine measurements are made at the beginning, 3rd, 6th and 12th weeks. Parathormone, vitamin D and calcium levels are measured before and after bone fracture for bone healing parameters. Bone healing and amount of callus formation is compared; biomechanically with three point bending test, radiologically with computed tomography and X-ray graphy, histopathologically and immunohistochemically with osteocalcine staining. Osteoblastic transformation of stem cells were assessed with DiI staining under immunoflorescent microscope. Group II, III ve IV rats with CKD had statistically significant weight loss. Morphological examination of callus formation and bone healing finding were as GroupIV> Group III> Group I> Group II. Biomechanical properties and resistance of bone tissue was measured as Group IV> Group III> Group I> Group II. Radiological findings are evaluated using modified Lane and Sandru scoring system which is a numerical scoring system that shows improvement as the score increases were resulted as; 3.91 ±0.64 (Group I), 3.5 ±0.64 (Group II), 5.08 ±1.44 (Group III), 5.43 ±0.99 (Group IV). The distance between cortical bone healing zone and outer margin of callus formation is measured as; 2.16 ±0.51 (Group I), 1.75 ±0.50 (Group II), 2.90 ±0.26 (Group III), 3.23 ±041 (Group IV). There was a correlation between results of bone healing scoring system and thickness of callus formation. Blood samples that were taken on first day, 3rd, 6th and 12th weeks show significant increase in plasma urea level and creatine and decrease in hemoglobin levels. All groups had decrease in calcium, parathormone and vitamin D levels. Rats with CKD had similar bone healing parameters comparing before bone fracture and after SVF and PRP treatment; decrease in vitamin and mineral levels had lower rate of decrease. Histopathologic evaluation was made using Allens scoring system which was resulted as; 4.71±0.99 (Group I), 4.17 ±0.62 (Group II), 5.17 ±0.90 (Group III), 5.38 ±0.88 (Group IV). Amount of bone mineralization was found as; 1.43 ±0.49 (Group I), 1.33 ±0.47 (Group II), 1.50 ±0.50 (Group III), 1.68 ±0.49 (Group IV). İmmunuhistochemical evaluation of bone tissue with osteocalcin staining resulted as; 1.43 ±0.49 (Group I), 1.33 ±0.47 (Group II), 2.17 ±0.37 (Group III), 2.38 ±0.49 (Group IV). Bone cells which are stained with osteocalcin were compared with slides with DiI staining. Transformation of adipose derived stem cells to bone cells were noted. In this study; We proved that negative effects of chronic renal disease on bone healing can be reduced by increasing mechanical, histological, radiological and biochemically with SVF and PRP treatments. Although thickness of callus tissue delays bone healing process, it also enhances biomechanical and organisation of bone tissue in long term. We conclude that stem cell therapies were effective for bone healing in animal models which can be promising for clinical trials.