Anat Cell Biol.  2024 Dec;57(4):559-569. 10.5115/acb.24.117.

Platelet-rich plasma protects hippocampal neurons and memory functions in a rat model of vascular dementia

• Moon JH
• Choi AL
• Noh HJ
• Song JH
• Hong GL
• Lee NS
• Jeong YG
• Han SY

• KMID: 2562503
• DOI: http://doi.org/10.5115/acb.24.117

Abstract

Platelet-rich plasma (PRP) is a promising biomaterial rich in bioactive growth factors, offering potential as a therapeutic agent for various diseases. However, its effectiveness in central nervous system disorders like vascular dementia (VaD) remains underexplored. This study investigated the potential of PRP to mitigate VaD progression in vivo. A rat model of VaD was established via bilateral common carotid artery occlusion and hypovolemia operation. Rats were randomly assigned to receive either PRP or platelet-poor plasma (PPP)—the latter being a byproduct of PRP preparation and used as a reference standard—resulting in the groups designated as ‘operated group (OP)+PRP’ and ‘OP+PPP’, respectively. PRP or PPP (500 μl) was administered intraperitoneally on the day of the operation and postoperative days 2, 4, 6, and 8. Cognitive function was assessed using the Y-maze, Barnes maze, and passive avoidance tests. On postoperative day 8, hippocampal samples were subjected to histological and semi-quantitative analyses. OP exhibited significant memory decline compared to controls, while the ‘OP+PRP’ group showed notable improvement. Histological analysis revealed increased neuronal loss and neuroinflammation in OP hippocampi, mitigated in ‘OP+PRP’. Semi-quantitative analysis showed decreased expression of brain-derived neurotrophic factor (BDNF) and its receptor tropomyosin receptor kinase B (TrkB) in OP, restored in ‘OP+PPP’ and further in ‘OP+PRP’. These results highlight PRP’s protective effects against VaD-induced hippocampal damage and cognitive impairment, partially attributed to BDNF/TrkB pathway upregulation.

Keyword

Platelet-rich plasma; Dementia; vascular; Brain-derived neurotrophic factor; Tropomyosin receptor kinase B

Figure

• 

  Fig. 1 Overview of the experimental protocol and Doppler flowmetry tracing in BCCAO/H operation. (A) Illustration of the preparation process for separating PPP and PRP from whole blood. The syringe diagram highlights the layers of blood components after centrifugation: PPP, PRP (containing buffy coat), and red blood cells. (B) Timeline showing the experimental schedule after BCCAO/H operation. Key assessments include the Y-MT, BMT, and PAT, alongside the time points for PRP or PPP administration (500 µl, intraperitoneally [i.p]). C-V staining and western blot analysis are planned following the behavioral tests. (C) Graph depicting the relative cerebral blood flow (rCBF) measured during the operation. The timeline includes significant events such as the Lt. CCA occlusion, Rt. CCA occlusion, and phases of blood withdrawal and reinfusion. Specific details such as the duration of the ischemic period (8 min marked) and the 20% value of rCBF baseline are shown to emphasize critical experimental manipulations. PPP, platelet-poor plasma; PRP, platelet-rich plasma; BCCAO/H, bilateral common carotid artery occlusion and hypovolemia; Y-MT, Y-maze test; BMT, Barnes maze test; PAT, passive avoidance test; C-V, cresyl violet; Lt. CCA, left common carotid artery; Rt. CCA, right common carotid artery.

• 

  Fig. 2 Y-maze test (Y-MT) results in platelet-rich plasma (PRP)-treated and untreated vascular dementia rat models. (A) Percentage of spontaneous alternation and (B) total number of arm entries in the Y-MT. Both graphs compare the Control group, Operation group (OP), operation group treated with platelet-poor plasma (OP+PPP), and operation group treated with PRP (OP+PRP). In both graphs, values are presented as mean±standard deviation (**P<0.01 vs. the Control group; #P<0.05 vs. the OP group).

• 

  Fig. 3 Barnes maze test (BMT) results in platelet-rich plasma (PRP)-treated and untreated vascular dementia rat models. (A) Representative tracking plots from the final day of trial test sessions in the BMT, illustrating the paths taken by rats in different groups. (B) Distance moved (cm) by rats over three trial days. (C) Latency (sec) to reach the goal platform across three trial days. (D) Detailed tracking plots highlighting the paths to the target quadrant and (E) the quantitative bar graphs of time spent in target quadrant obtained from probe test sessions in the BMT. In (D), the location where the escape box was situated during the trial test session is depicted as a dotted red circle. All tests compare the Control group, Operation group (OP), operation group treated with platelet-poor plasma (OP+PPP), and operation group treated with PRP (OP+PRP). In all graphs, values are presented as mean±standard deviation (***P<0.001 vs. the Control group; #P<0.05, ##P<0.01, and ###P<0.001 vs. the OP group).

• 

  Fig. 4 Passive avoidance test (PAT) results in platelet-rich plasma (PRP)-treated and untreated vascular dementia rat models. (A) Step-through latencies during the test session and (B) those during the training session. Both tests compare the Control group, Operation group (OP), operation group treated with platelet-poor plasma (OP+PPP), and operation group treated with PRP (OP+PRP). In both graphs, values are presented as mean±standard deviation (***P<0.001 vs. the Control group; ###P<0.001 vs. the OP group).

• 

  Fig. 5 Histological assessment of neuronal viability and inflammation in hippocampi of platelet-rich plasma (PRP)-treated and untreated vascular dementia rat models. (A) Representative images of hippocampal sections stained to highlight neuronal viability and inflammation. Upper panels show the overall hippocampal structure with regions of interest (ROI) marked in red boxes. Lower panels are magnified views from the hippocampal CA1 region, indicating viable neurons (red arrowheads) and cells indicative of infiltrating inflammatory cells and activated glial cells, used as a surrogate of neuroinflammation (green arrowheads). Scale bar=500 µm. (B) Quantitative bar graphs of viable neurons and (C) neuroinflammation observed in the ROI. These assessments compare the Control group, Operation group (OP), operation group treated with platelet-poor plasma (OP+PPP), and operation group treated with PRP (OP+PRP). In graphs (B) and (C), values are presented as mean±standard deviation (***P<0.001 vs. the Control group; ###P<0.001 vs. the OP group). CA, cornu ammonis; DG, dentate gyrus.

• 

  Fig. 6 Western blot results of the TrkB/BDNF signaling pathway using hippocampal homogenates from PRP-treated and untreated vascular dementia rat models. (A) Representative western blot band images and quantitative graphs of (B) TrkB and (C) BDNF. The analysis compares the Control group, Operation group (OP), operation group treated with platelet-poor plasma (OP+PPP), and operation group treated with platelet-rich plasma (OP+PRP). In graphs (B) and (C), values were normalized by β-actin. Data are presented as mean±standard deviation (*P<0.05 and **P<0.01 vs. the Control group; ##P<0.01 and ###P<0.001 vs. the OP group). TrkB, tropomyosin receptor kinase B; BDNF, brain-derived neurotrophic factor.

Reference

References

1. Venkat P, Chopp M, Chen J. 2015; Models and mechanisms of vascular dementia. Exp Neurol. 272:97–108. DOI: 10.1016/j.expneurol.2015.05.006. PMID: 25987538. PMCID: PMC4631710.

2. Sengupta P, Ganguly J, Pal S, Ghosal M. 2019; Pattern of cognitive deficits in vascular dementia. Indian J Med Res. 149:503–7. DOI: 10.4103/ijmr.IJMR_1336_17. PMID: 31411174. PMCID: PMC6676860.

3. Dichgans M, Leys D. 2017; Vascular cognitive impairment. Circ Res. 120:573–91. DOI: 10.1161/CIRCRESAHA.116.308426. PMID: 28154105.

4. Kharkongor R, Nambi P, Radhakrishnan R. 2024; Fucoidan protects CA1 pyramidal neurons of the hippocampus and preserves the cognitive profile of rats subjected to transient forebrain ischemia. Brain Res. 1828:148769. DOI: 10.1016/j.brainres.2024.148769. PMID: 38237671.

5. Bir SC, Khan MW, Javalkar V, Toledo EG, Kelley RE. 2021; Emerging concepts in vascular dementia: a review. J Stroke Cerebrovasc Dis. 30:105864. DOI: 10.1016/j.jstrokecerebrovasdis.2021.105864. PMID: 34062312.

6. Patil P, Jadhav M, Suvvari TK, Thomas V. 2024; Therapeutic uses of platelet-rich plasma (PRP) in sport injuries - a narrative review. J Orthop Rep. 3:100287. DOI: 10.1016/j.jorep.2023.100287.

7. Verma R, Kumar S, Garg P, Verma YK. 2023; Platelet-rich plasma: a comparative and economical therapy for wound healing and tissue regeneration. Cell Tissue Bank. 24:285–306. DOI: 10.1007/s10561-022-10039-z. PMID: 36222966. PMCID: PMC9555256.

8. Borhani-Haghighi M, Mohamadi Y. 2019; The therapeutic effect of platelet-rich plasma on the experimental autoimmune encephalomyelitis mice. J Neuroimmunol. 333:476958. DOI: 10.1016/j.jneuroim.2019.04.018. PMID: 31103935.

9. Simonian NA, Coyle JT. 1996; Oxidative stress in neurodegenerative diseases. Annu Rev Pharmacol Toxicol. 36:83–106. DOI: 10.1146/annurev.pa.36.040196.000503. PMID: 8725383.

10. Nunomura A, Perry G, Aliev G, Hirai K, Takeda A, Balraj EK, Jones PK, Ghanbari H, Wataya T, Shimohama S, Chiba S, Atwood CS, Petersen RB, Smith MA. 2001; Oxidative damage is the earliest event in Alzheimer disease. J Neuropathol Exp Neurol. 60:759–67. DOI: 10.1093/jnen/60.8.759. PMID: 11487050.

11. Zhang J, Zhang J, Zhang N, Li T, Zhou X, Jia J, Liang Y, Sun X, Chen H. 2020; The effects of platelet-rich and platelet-poor plasma on biological characteristics of BM-MSCs in vitro. Anal Cell Pathol (Amst). 2020:8546231. DOI: 10.1155/2020/8546231. PMID: 32908815. PMCID: PMC7471809.

12. National Research Council (US) Committee for the Update of the Guide for the Care and Use of Laboratory Animals. Guide for the care and use of laboratory animals. 8th ed. National Academies Press (US);2011.

13. Dhurat R, Sukesh M. 2014; Principles and methods of preparation of platelet-rich plasma: a review and author's perspective. J Cutan Aesthet Surg. 7:189–97. DOI: 10.4103/0974-2077.150734. PMID: 25722595. PMCID: PMC4338460.

14. Kang HB, Kim SH, Uhm SH, Kim DK, Lee NS, Jeong YG, Sung NY, Kim DS, Han IJ, Yoo YC, Han SY. 2022; Perilla frutescens leaf extract attenuates vascular dementia-associated memory deficits, neuronal damages, and microglial activation. Curr Issues Mol Biol. 44:257–72. DOI: 10.3390/cimb44010019. PMID: 35723398. PMCID: PMC8929086.

15. Sung NS, Uhm SH, Kang HB, Lee NS, Jeong YG, Kim DK, Sung NY, Kim DS, Yoo YC, Han SY. 2023; Rubus fruticosus leaf extract inhibits vascular dementia-induced memory impairment and neuronal loss by attenuating neuroinflammation. Anat Cell Biol. 56:494–507. DOI: 10.5115/acb.23.195. PMID: 37743615. PMCID: PMC10714077.

16. Alshammari TK, Alghamdi H, Green TA, Niazy A, Alkahdar L, Alrasheed N, Alhosaini K, Alswayyed M, Elango R, Laezza F, Alshammari MA, Yacoub H. 2019; Assessing the role of toll-like receptor in isolated, standard and enriched housing conditions. PLoS One. 14:e0222818. DOI: 10.1371/journal.pone.0222818. PMID: 31647818. PMCID: PMC6812767.

17. Graham NL, Emery T, Hodges JR. 2004; Distinctive cognitive profiles in Alzheimer's disease and subcortical vascular dementia. J Neurol Neurosurg Psychiatry. 75:61–71.

18. Choi DH, Lee KH, Kim JH, Seo JH, Kim HY, Shin CY, Han JS, Han SH, Kim YS, Lee J. 2014; NADPH oxidase 1, a novel molecular source of ROS in hippocampal neuronal death in vascular dementia. Antioxid Redox Signal. 21:533–50. DOI: 10.1089/ars.2012.5129. PMID: 24294978. PMCID: PMC4086030.

19. Shcherbitskaia AD, Vasilev DS, Milyutina YP, Tumanova NL, Mikhel AV, Zalozniaia IV, Arutjunyan AV. 2021; Prenatal hyperhomocysteinemia induces glial activation and alters neuroinflammatory marker expression in infant rat hippocampus. Cells. 10:1536. DOI: 10.3390/cells10061536. PMID: 34207057. PMCID: PMC8234222.

20. Huang Y, Li Z, Nan G. 2017; Effect of hippocampal L-NBP on BDNF and TrkB expression and neurological function of vascular dementia rats. Mol Med Rep. 16:7673–8. DOI: 10.3892/mmr.2017.7539. PMID: 28944854.

21. Yin C, Deng Y, Liu Y, Gao J, Yan L, Gong Q. 2018; Icariside II ameliorates cognitive impairments induced by chronic cerebral hypoperfusion by inhibiting the amyloidogenic pathway: involvement of BDNF/TrkB/CREB signaling and up-regulation of PPARα and PPARγ in rats. Front Pharmacol. 9:1211. DOI: 10.3389/fphar.2018.01211. PMID: 30405422. PMCID: PMC6206175.

22. Kuter DJ, Rummel M, Boccia R, Macik BG, Pabinger I, Selleslag D, Rodeghiero F, Chong BH, Wang X, Berger DP. 2010; Romiplostim or standard of care in patients with immune thrombocytopenia. N Engl J Med. 363:1889–99. DOI: 10.1056/NEJMoa1002625. PMID: 21067381.

23. Oneto P, Etulain J. 2021; PRP in wound healing applications. Platelets. 32:189–99. DOI: 10.1080/09537104.2020.1849605. PMID: 33251921.

24. Foster TE, Puskas BL, Mandelbaum BR, Gerhardt MB, Rodeo SA. 2009; Platelet-rich plasma: from basic science to clinical applications. Am J Sports Med. 37:2259–72. DOI: 10.1177/0363546509349921. PMID: 19875361.

25. Mazzucco L, Balbo V, Cattana E, Guaschino R, Borzini P. 2009; Not every PRP-gel is born equal. Evaluation of growth factor availability for tissues through four PRP-gel preparations: Fibrinet, RegenPRP-Kit, Plateltex and one manual procedure. Vox Sang. 97:110–8. DOI: 10.1111/j.1423-0410.2009.01188.x. PMID: 19392780.

26. Anitua E, Andia I, Ardanza B, Nurden P, Nurden AT. 2004; Autologous platelets as a source of proteins for healing and tissue regeneration. Thromb Haemost. 91:4–15. DOI: 10.1160/TH03-07-0440. PMID: 14691563.

27. Giusti I, Rughetti A, D'Ascenzo S, Millimaggi D, Pavan A, Dell'Orso L, Dolo V. 2009; Identification of an optimal concentration of platelet gel for promoting angiogenesis in human endothelial cells. Transfusion. 49:771–8. DOI: 10.1111/j.1537-2995.2008.02033.x. PMID: 19170984.

28. Linker SB, Mendes APD, Marchetto MC. 2020; IGF-1 treatment causes unique transcriptional response in neurons from individuals with idiopathic autism. Mol Autism. 11:55. DOI: 10.1186/s13229-020-00359-w. PMID: 32591005. PMCID: PMC7320548.

29. Silva-Hucha S, Pastor AM, Morcuende S. 2021; Neuroprotective effect of vascular endothelial growth factor on motoneurons of the oculomotor system. Int J Mol Sci. 22:814. DOI: 10.3390/ijms22020814. PMID: 33467517. PMCID: PMC7830098.

30. Paul G, Zachrisson O, Varrone A, Almqvist P, Jerling M, Lind G, Rehncrona S, Linderoth B, Bjartmarz H, Shafer LL, Coffey R, Svensson M, Mercer KJ, Forsberg A, Halldin C, Svenningsson P, Widner H, Frisén J, Pålhagen S, Haegerstrand A. 2015; Safety and tolerability of intracerebroventricular PDGF-BB in Parkinson's disease patients. J Clin Invest. 125:1339–46. DOI: 10.1172/JCI79635. PMID: 25689258. PMCID: PMC4362250.

31. Li H, Gan X, Pan L, Zhang Y, Hu X, Wang Z. 2022; EGF/bFGF promotes survival, migration and differentiation into neurons of GFP-labeled rhesus monkey neural stem cells xenografted into the rat brain. Biochem Biophys Res Commun. 620:76–82. DOI: 10.1016/j.bbrc.2022.06.077. PMID: 35780584.

32. Nolte M, Margadant C. 2020; Controlling immunity and inflammation through integrin-dependent regulation of TGF-β. Trends Cell Biol. 30:49–59. DOI: 10.1016/j.tcb.2019.10.002. PMID: 31744661.

33. Kandasamy M, Anusuyadevi M, Aigner KM, Unger MS, Kniewallner KM, de Sousa DMB, Altendorfer B, Mrowetz H, Bogdahn U, Aigner L. 2020; TGF-β signaling: a therapeutic target to reinstate regenerative plasticity in vascular dementia? Aging Dis. 11:828–50. DOI: 10.14336/AD.2020.0222. PMID: 32765949. PMCID: PMC7390515.

34. Nie PY, Ji LL, Fu CH, Peng JB, Wang ZY, Tong L. 2021; miR-132 regulates PTSD-like behaviors in rats following single-prolonged stress through fragile X-related protein 1. Cell Mol Neurobiol. 41:327–40. DOI: 10.1007/s10571-020-00854-x. PMID: 32333305.

35. Jeelani M. 2024; miRNAs in epilepsy: a review from molecular signatures to therapeutic intervention. Int J Biol Macromol. 263(Pt 2):130468. DOI: 10.1016/j.ijbiomac.2024.130468. PMID: 38417757.

36. Le Blanc J, Fleury S, Boukhatem I, Bélanger JC, Welman M, Lordkipanidzé M. 2020; Platelets selectively regulate the release of BDNF, but not that of its precursor protein, proBDNF. Front Immunol. 11:575607. DOI: 10.3389/fimmu.2020.575607. PMID: 33324399. PMCID: PMC7723927.

37. Cheng PL, Song AH, Wong YH, Wang S, Zhang X, Poo MM. 2011; Self-amplifying autocrine actions of BDNF in axon development. Proc Natl Acad Sci U S A. 108:18430–5. DOI: 10.1073/pnas.1115907108. PMID: 22025720. PMCID: PMC3215064.

38. Shelly M, Cancedda L, Lim BK, Popescu AT, Cheng PL, Gao H, Poo MM. 2011; Semaphorin3A regulates neuronal polarization by suppressing axon formation and promoting dendrite growth. Neuron. 71:433–46. DOI: 10.1016/j.neuron.2011.06.041. PMID: 21835341. PMCID: PMC3164872.

39. Pound P, Ritskes-Hoitinga M. 2018; Is it possible to overcome issues of external validity in preclinical animal research? Why most animal models are bound to fail. J Transl Med. 16:304. DOI: 10.1186/s12967-018-1678-1. PMID: 30404629. PMCID: PMC6223056.