Begin typing your search above and press return to search.
Volume: 21 Issue: 1 January 2023

FULL TEXT

ARTICLE
Positive Effect of Platelet-Rich Plasma on Neoureterocystostomy in Rats: An Experimental Study

Objectives: Some of the most common problems after kidney transplant are urologic complications, including ureterocystoanastomosis leakage and stenosis with the development of severe renal graft complications. Isolated plasma contains active substances that cause the activation of various growth factors for the processes of tissue repair or regeneration, has an antiinflam-matory effect, activates angiogenesis, and reduces the risk of infectious complications. Platelet-rich plasma is actively used to stimulate bone regeneration, heal wounds and ulcers on the skin, enhance reconstruction of the larynx and trachea, and ameliorate urethral damage, among other uses. This study was developed to evaluate the positive effect of platelet-rich plasma on the healing process of an anastomotic wound in a model of ureterocystoanas-tomosis in rats.
Material and Methods: We randomized 14 Wistar albino male rats into 2 groups: group 1 included 7 rats that received platelet-rich plasma after ureterocystostomy; group 2 was the control group and included 7 rats that underwent ureterocystostomy without platelet-rich plasma. On postoperative day 7, all animals were euthanized, and the anastomosis area was resected for determination of the tissue hydroxyproline levels and histopathology examination.
Results: Tissue hydroxyproline levels were 767 ± 62.9 ?g/g in group 1 and 256 ± 28.0 ?g/g in group 2. Tissue hydroxyproline levels were significantly higher in group 1 compared with group 2 (P < .05). There were no significant differences in epithelial damage, acute inflammation, or fibrosis between the tissue samples of both groups.
Conclusions: The results of this study show that the use of platelet-rich plasma during ureterocystostomy produces a positive effect. Our further research will be devoted to the use of autologous platelet-rich plasma in ureterocystostomy in big models.


Key words : Kidney transplant, Ureterocystoanastomosis, Ureterocystostomy, Ureteroneocystostomy, Urologic complications

Introduction

Some of the most common problems after kidney transplant are urologic complications, which occur in 2.5% to 30.0% of cases.1,2 Urine leakage is the most common early urologic complication after renal transplant, with an incidence of 1.2% to 8.9%.3 Early urologic complications include neoureterocysto-anastomosis (NUCA) leakage and stenosis with the development of severe complications, and a strategy is required for the prevention of these complications.4,5 The NUCA leakage can be caused by insufficient blood supply to the donor ureter, and damage of the so-called golden triangle structures can lead to necrosis of the ureteral distal portion in up to 70% of cases.2,5,6

Isolated plasma contains active substances that cause the activation of various growth factors for the processes of tissue repair or regeneration, has an anti-inflammatory effect, activates local angiogenesis, and reduces the risk of local infectious complications.7-9

Platelet-rich plasma (PRP) is actively used for the following purposes: to stimulate bone regeneration, heal wounds and skin ulcers, enhance the reconstruction of the larynx and trachea, treat musculoskeletal and nerve injuries, promote the engraftment of various types of skin flaps, support gastrointestinal anastomoses, and ameliorate urethral damage, among other purposes.10-16

This study was developed to evaluate the positive effect of PRP on the healing process of an anasto-motic wound in a model of ureterocystostomy (UCS) in rats.

Materials and Methods

Animals and ethics statement

We randomized 14 Wistar albino male rats, weighing 360 to 536 g, into 2 groups: group 1 consisted of 7 rats that received PRP during UCS; group 2 was the control group of 7 rats that underwent UCS without PRP (Table 1). The rats of both groups were kept in the same conditions before and after the surgery, in compliance with the necessary regimens for temperature, light, and food. This study was approved by the Baskent University Ethical Committee for Experimental Research on Animals (project No. E-94603339-604.01.02-38157).

Platelet-rich plasma preparation

Blood was obtained by whole body blood collection from 3 donor rats with ketamine anesthesia (60 mg/kg; VetaKetam, Vet-Agro) through cardiac puncture. A volume of 58 mL of blood contained 276 × 109 platelets/L. All collected blood was placed into tubes with 3.8% sodium citrate. Tubes were centrifuged twice, as described by Tobita and colleagues: 10 minutes at 2400 rpm and 15 minutes at 3600 rpm.17 After the first centrifugation, a long needle was used to collect the middle layer with leukocytes and thrombocytes and transfer into another tube. After the second centrifugation, the platelet count was 942 × 109 platelets/L, which was 3.4-fold greater than the initial amount. Before PRP in group 1 was used, plasma with concentrated platelets was activated by 10% calcium chloride solution, and a gel-like mass was obtained (Figure 1).

Surgical procedure

All the rats in each group underwent surgery on the same day and by the same surgeon for technical uniformity. After an overnight fast, the rats were anesthetized by intramuscular injection of ketamine (60 mg/kg; VetaKetam) and xylazine (10 mg/kg; VetaXyl, Vet-Agro). The rats were on spontaneous respiration during the surgery. The abdominal wall of each rat was shaved; under aseptic conditions, a 4-cm incision was made along the midline of the abdomen. In both groups, the left ureter was crossed at the level of the confluence with the bladder, and then an anastomosis was performed with single-layer interrupted 8-0 absorbable sutures. In group 1, activated PRP gel was applied to the ureterocys-toanastomosis line. The laparotomy incision was closed with a continuous absorbable 3-0 suture (Figure 2). Postoperative conditions were the same in both groups, with feeding started the next day after surgery. No postoperative analgesics or antibiotics were use.

On postoperative day 7, all animals were exsanguinated through percutaneous cardiac puncture under ketamine anesthesia by the method described by Donovan and Brown.18 The rat abdominal wound was reopened, and the anastomosis zone was carefully dissected and the anastomosis was located. The anastomosis area was resected with a margin of at least 2 cm proximally and distally for determination of the tissue hydroxyproline (THP) levels and histopathology examination.

Tissue hydroxyproline measurement

The excised and purified anastomosis line tissue was cooled, neutralized in potassium hydroxide, and oxidized with alanine and potassium borate buffers with the addition of potassium chloride. A solution of chloramine-T, thiosulfate, and toluene was added and left at room temperature for 20 minutes. After the sample was stained with hematoxylin-eosin, the color and intensity were evaluated spectrophoto-metrically at a wavelength of 560 nm.

Histopathology analysis

For histopathology examination of the resected anastomosis area, tissue was fixed in 10% formalin and embedded in paraffin, and 4-?m sections were stained with hematoxylin-eosin. Epithelial damage, acute inflammation, and fibrosis were determined as parameters of inflammation in the tissue samples. Each parameter was evaluated on a scale of 0 to 3 points: 0 = absence, 1 = mild, 2 = moderate, and 3 = severe.

Statistical analyses

We used SPSS software (version 20.0 for Windows) for statistical analyses; the Mann-Whitney U test was used to compare the variables of the 2 groups with median values. Statistical differences between the 2 groups were considered significant at values of P < .05.

Results

The levels of THP in both groups are shown in Table 2. The mean THP levels were 767 ± 62.9 ?g/g in group 1 and 256 ± 28.0 ?g/g in group 2 (control), and this difference was statistically significant (P < .05). These data indicated a positive effect of PRP application on the NUCA.

The results of the histology evaluations are shown in Table 3. There were no significant differences in epithelial damage, acute inflammation, or fibrosis between the tissue samples of group 1 and group 2 (Figure 3).

Discussion

Ureteral anastomosis leakage remains a serious problem, especially in kidney transplant, and is associated with a high risk of graft failure and recipient mortality.2,19 Previous studies have investigated and described the risk factors that can affect the healing of the UCS.20,21 To date, various methods of UCS are in use for kidney transplantation, the most common of which is the Lich-Gregoir method. Some authors claim that the Lich-Gregoir method has an advantage versus other methods by preventing urologic complications in UCS.22 In a similar manner, Szabo-Pap and colleagues did not find a significant difference between the types of urinary anastomosis and the frequency of leakage in kidney transplant recipients.23 Also, Peng and colleagues have reported that there is no evidence that ureteral stents could prevent the occurrence of ureteral anastomosis leakage.24

The local use of PRP has been mostly limited to studies of intestinal anastomoses and the positive effect of PRP in the healing of intestinal anastomoses.15,25-28 In the available literature, we did not find any report of local use of PRP to strengthen the urinary or ureteral anastomosis. We found a single experimental study by Tavukcu and colleagues on the use of PRP in urethral trauma.16

We used double centrifugation in the preparation of PRP, which facilitated an increase in the initial platelet concentration by 3.4-fold, to 942 × 109 platelets/L, and this corresponds to the recommen-dations of Tambella and colleagues.29 In our study, we euthanized the rats and obtained tissue samples for examination on postoperative day 7. Usually in clinical practice, anastomosis leakages are diagnosed during the period from postoperative day 5 to day 7.30 After euthanasia, we did not find any signs of anastomosis leakage or ureteral dilation in the surgical field in the rats.

In this study, we found no differences between the 2 groups in terms of histology parameters. However, THP levels in the PRP group showed significantly better results than in the control group.

There were some limitations in our study. We used homologous blood from 3 donor rats for PRP preparation. Theoretically, these homologous blood cells could cause the processes of immunologic inflammation in the area of application of PRP. Another limitation is the reimplant of the ureter without prior nephroureterectomy. That is, we did not use the kidney transplant model in these rats, which undoubtedly carries risk factors against the healing of urinary anastomosis in clinical practice. To exclude such influences, in the future we plan to use autologous PRP for ureteral anastomoses in big models.

In conclusion, the results of this study showed that the use of PRP during UCS could have a positive effect. Platelet-rich plasma has potential usefulness to prevent leakage of ureterocystoanastomosis after kidney transplant. To confirm this fact, further studies with big experimental models and the use of autologous PRP are necessary.


References:

  1. Dalgic A, Boyvat F, Karakayali H, Moray G, Emiroglu R, Haberal M. Urologic complications in 1523 renal transplantations: the Baskent University experience. Transplant Proc. 2006;38(2):543-547. doi:10.1016/j.transproceed.2005.12.116
    CrossRef - PubMed
  2. Haberal M, Boyvat F, Akdur A, Kirnap M, Ozcelik U, Karakayali FY. Surgical complications after kidney transplantation. Exp Clin Transplant. 2016;14(6):587-595. doi:10.6002/ect.2016.0290
    CrossRef - PubMed
  3. Bruintjes MHD, d’Ancona FCH, Zhu X, Hoitsma AJ, Warle MC. An update on early urological complications in kidney transplantation: a national cohort study. Ann Transplant. 2019;24:617-624. doi:10.12659/AOT.920086
    CrossRef - PubMed
  4. Duty BD, Barry JM. Diagnosis and management of ureteral complications following renal transplantation. Asian J Urol. 2015;2(4):202-207. doi:10.1016/j.ajur.2015.08.002
    CrossRef - PubMed
  5. Praz V, Leisinger HJ, Pascual M, Jichlinski P. Urological complications in renal transplantation from cadaveric donor grafts: a retrospective analysis of 20 years. Urol Int. 2005;75(2):144-149. doi:10.1159/000087169
    CrossRef - PubMed
  6. Buttigieg J, Agius-Anastasi A, Sharma A, Halawa A. Early urological complications after kidney transplantation: an overview. World J Transplant. 2018;8(5):142-149. doi:10.5500/wjt.v8.i5.142
    CrossRef - PubMed
  7. Mohan SP, Jaishangar N, Devy S, Narayanan A, Cherian D, Madhavan SS. Platelet-rich plasma and platelet-rich fibrin in periodontal regeneration: a review. J Pharm Bioallied Sci. 2019;11(Suppl 2):S126-S130. doi:10.4103/JPBS.JPBS_41_19
    CrossRef - PubMed
  8. Nasirzade J, Kargarpour Z, Hasannia S, Strauss FJ, Gruber R. Platelet-rich fibrin elicits an anti-inflammatory response in macrophages in vitro. J Periodontol. 2020;91(2):244-252. doi:10.1002/JPER.19-0216
    CrossRef - PubMed
  9. Cieslik-Bielecka A, Bold T, Ziolkowski G, Pierchala M, Krolikowska A, Reichert P. Antibacterial activity of leukocyte- and platelet-rich plasma: an in vitro study. Biomed Res Int. 2018;2018:9471723. doi:10.1155/2018/9471723
    CrossRef - PubMed
  10. Oryan A, Alidadi S, Moshiri A. Platelet-rich plasma for bone healing and regeneration. Expert Opin Biol Ther. 2016;16(2):213-232. doi:10.1517/14712598.2016.1118458
    CrossRef - PubMed
  11. Milek T, Nagraba L, Mitek T, et al. Autologous platelet-rich plasma reduces healing time of chronic venous leg ulcers: a prospective observational study. Adv Exp Med Biol. 2019;1176:109-117. doi:10.1007/5584_2019_388
    CrossRef - PubMed
  12. Sahin Onder S, Sahin Yilmaz A, Erkmen B, Topal CS, Gergin O, Canpolat MS. Platelet-rich plasma for laryngotracheal reconstruction: an experimental study. Eur Arch Otorhinolaryngol. 2020;277(11):3103-3109. doi:10.1007/s00405-020-06091-8
    CrossRef - PubMed
  13. Kucuk L, Gunay H, Erbas O, Kucuk U, Atamaz F, Coskunol E. Effects of platelet-rich plasma on nerve regeneration in a rat model. Acta Orthop Traumatol Turc. 2014;48(4):449-454. doi:10.3944/AOTT.2014.13.0029
    CrossRef - PubMed
  14. Slaninka I, Fibir A, Kaska M, Paral J. Use of autologous platelet-rich plasma in healing skin graft donor sites. J Wound Care. 2020;29(1):36-41. doi:10.12968/jowc.2020.29.1.36
    CrossRef - PubMed
  15. Sozutek A, Colak T, Cetinkunar S, et al. The effect of platelet-rich-plasma on the healing of left colonic anastomosis in a rat model of intra-abdominal sepsis. J Invest Surg. 2016;29(5):294-301. doi:10.3109/08941939.2015.1111473
    CrossRef - PubMed
  16. Tavukcu HH, Aytac O, Atug F, et al. Protective effect of platelet-rich plasma on urethral injury model of male rats. Neurourol Urodyn. 2018;37(4):1286-1293. doi:10.1002/nau.23460
    CrossRef - PubMed
  17. Tobita M, Uysal AC, Ogawa R, Hyakusoku H, Mizuno H. Periodontal tissue regeneration with adipose-derived stem cells. Tissue Eng Part A. 2008;14(6):945-953. doi:10.1089/ten.tea.2007.0048
    CrossRef - PubMed
  18. Donovan J, Brown P. Euthanasia. Curr Protoc Immunol. 2006;73:1.8.1-1.8.4. doi:10.1002/0471142735.im0108s73
    CrossRef - PubMed
  19. Choi YS, Kim KS, Choi SW, et al. Ureteral complications in kidney transplantation: analysis and management of 853 consecutive laparoscopic living-donor nephrectomies in a single center. Transplant Proc. 2016;48(8):2684-2688. doi:10.1016/j.transproceed.2016.06.054
    CrossRef - PubMed
  20. Jenjitranant P, Tansakul P, Sirisreetreerux P, Leenanupunth C, Jirasiritham S. Risk factors for anastomosis leakage after kidney transplantation. Res Rep Urol. 2020;12:509-516. doi:10.2147/RRU.S272899
    CrossRef - PubMed
  21. Pillot P, Bardonnaud N, Lillaz J, et al. Risk factors for surgical complications after renal transplantation and impact on patient and graft survival. Transplant Proc. 2012;44(9):2803-2808. doi:10.1016/j.transproceed.2012.09.030
    CrossRef - PubMed
  22. Alberts VP, Idu MM, Legemate DA, Laguna Pes MP, Minnee RC. Ureterovesical anastomotic techniques for kidney transplantation: a systematic review and meta-analysis. Transpl Int. 2014;27(6):593-605. doi:10.1111/tri.12301
    CrossRef - PubMed
  23. Szabo-Pap M, Zadori G, Fedor R, et al. Surgical complications following kidney transplantations: a single-center study in Hungary. Transplant Proc. 2016;48(7):2548-2551. doi:10.1016/j.transproceed.2016.07.012
    CrossRef - PubMed
  24. Peng YL, Ning K, Wu ZS, et al. Ureteral stents cannot decrease the incidence of ureteroileal anastomotic stricture and leakage: a systematic review and meta-analysis. Int J Surg. 2021;93:106058. doi:10.1016/j.ijsu.2021.106058
    CrossRef - PubMed
  25. Yamaguchi R, Terashima H, Yoneyama S, Tadano S, Ohkohchi N. Effects of platelet-rich plasma on intestinal anastomotic healing in rats: PRP concentration is a key factor. J Surg Res. 2012;173(2):258-266. doi:10.1016/j.jss.2010.10.001
    CrossRef - PubMed
  26. Zhou B, Ren J, Ding C, et al. Protection of colonic anastomosis with platelet-rich plasma gel in the open abdomen. Injury. 2014;45(5):864-868. doi:10.1016/j.injury.2014.01.018
    CrossRef - PubMed
  27. Daradka M, Alardah MM, Ismail ZB. Effects of autologous platelet-rich plasma coated sutures on intestinal anastomotic healing in rabbits. Heliyon. 2019;5(11):e02713. doi:10.1016/j.heliyon.2019.e02713
    CrossRef - PubMed
  28. Aydin MA, Guler EM, Demiroz AS, Aydin MF, Saglam G. Comparison of platelet-rich plasma-impregnated suture material with low and high platelet concentration to improve colonic anastomotic wound healing in rats. Gastroenterol Res Pract. 2020;2020:7386285. doi:10.1155/2020/7386285
    CrossRef - PubMed
  29. Tambella AM, Attili AR, Dupre G, et al. Platelet-rich plasma to treat experimentally-induced skin wounds in animals: a systematic review and meta-analysis. PLoS One. 2018;13(1):e0191093. doi:10.1371/journal.pone.0191093
    CrossRef - PubMed
  30. Daglioglu YK, Duzgun O, Sarici IS, Ulutas KT. Comparison of platelet rich plasma versus fibrin glue on colonic anastomoses in rats. Acta Cir Bras. 2018;33(4):333-340. doi:10.1590/s0102-865020180040000005
    CrossRef - PubMed


Volume : 21
Issue : 1
Pages : 47 - 51
DOI : 10.6002/ect.2022.0333


PDF VIEW [958] KB.
FULL PDF VIEW

From the 1Department of Surgery 2, West-Kazakhstan Medical University, and the 2Department of Surgery and Transplantation, Aktobe Medical Center, Aktobe, Kazakhstan; the 3Department of General Surgery, Division of Transplantation, Baskent University; and the 4Department of Medical Pathology, Faculty of Medicine, Ankara, Turkey
Acknowledgements: The authors have not received any funding or grants in support of the presented research or for the preparation of this work and have no declarations of potential conflicts of interest.
Corresponding author: Myltykbay Rysmakhanov. Department of Surgery 2, West-Kazakhstan Medical University, 68, Maresyev Street, Aktobe city, Kazakhstan, 040017
E-mail: mrtransplantolog@gmail.com