Comparative analysis of the regenerative potential of blood derivatives on a cell model of corneal stromal injury

Abstract

BACKGROUND: Agents based on blood derivatives contain substances that accelerate the healing of wound defects. Nowadays data have been accumulated that various blood derivatives have different effects on regeneration processes. Therefore, a comparative study of the effect of several types of such preparations on the corneal stroma’s cell culture is relevant.

AIM: Impact assessment of 3 different blood derivatives on the processes of proliferation, migration and cell death of keratocytes in vitro.

MATERIAL AND METHODS: This study was conducted on a primary cell culture of human corneal keratocytes. Blood samples were obtained from the cubital vein of healthy volunteers after signing informed consent to participate in the study. Monolayer wound healing test, formazan test to assess proliferation, staining for Annexin V to assess the processes of apoptosis and necrosis were perfomed. Moreover, effect evaluation of platelet-rich plasma, intact serum and processed serum were performed.

RESULTS: It was shown that all 3 stimulants have a positive effect on the regeneration of the experimental stromal defect, but it is implemented in different ways. Processed whey showed the greatest stimulating effect on the process of cell migration. This serum reduced the level of cell death, but had almost no effect on proliferation processes. It is advisable to further study different types of stimulants to obtain processed serum. Intact serum increased the percentage of dividing cells more than others, but also proportionally increased the percentage of death in culture. Platelet-rich plasma showed the weakest effect on the migratory ability of keratocytes, but it still differed significantly from the control. This stimulator had little effect on the process of cell division, and had no effect on the overall level of cell death.

CONCLUSION: Experimental data can be taken into account in choosing of the management in clinical practice and personalized usage of blood derivatives, specifically in prescribing some blood derivatives to accelerate regeneration, and other derivatives for reducing cell death’s processes.

Keywords

corneal stroma / keratocytes / serum / polyA:U / platelet-rich plasma / cell culture / growth factors

References

[1] Wang EY, Kong X, Wolle M, et al. Global trends in blindness and vision impairment resulting from corneal opacity 1984-2020: a meta-analysis. Ophthalmology. 2023:S0161-6420(23)00187-2. doi: 10.1016/j.ophtha.2023.03.012

[2] Wang E.Y., Kong X., Wolle M., et al. Global trends in blindness and vision impairment resulting from corneal opacity 1984-2020: a meta-analysis // Ophthalmology. 2023. P. S0161-6420(23)00187-2. doi: 10.1016/j.ophtha.2023.03.012

[3] Ung L, Bispo P, Shanbhag S, et al. The persistent dilemma of microbial keratitis: Global burden, diagnosis, and antimicrobial resistance. Surv Ophthalmol. 2019;64(3):255–271. doi: 10.1016/j.survophthal.2018.12.003

[4] Ung L., Bispo P., Shanbhag S., et al. The persistent dilemma of microbial keratitis: Global burden, diagnosis, and antimicrobial resistance // Surv Ophthalmol. 2019. Vol. 64, N 3. P. 255–271. doi: 10.1016/j.survophthal.2018.12.003

[5] Ting D, Ho C, Cairns J, et al. 12-year analysis of incidence, microbiological profiles and in vitro antimicrobial susceptibility of infectious keratitis: the Nottingham Infectious Keratitis Study. Br J Ophthalmol. 2020;105(3):328–333. doi: 10.1136/bjophthalmol-2020-316128

[6] Ting D., Ho C., Cairns J. et al. 12-year analysis of incidence, microbiological profiles and in vitro antimicrobial susceptibility of infectious keratitis: the Nottingham Infectious Keratitis Study // Br J Ophthalmol. 2020. Vol. 105, N 3. P. 328–333. doi: 10.1136/bjophthalmol-2020-316128

[7] Ligocki AJ, Fury W, Gutierrez C, et al. Molecular characteristics and spatial distribution of adult human corneal cell subtypes. Sci Rep. 2021;11(1):16323. doi: 10.1038/s41598-021-94933-8

[8] Ligocki A.J., Fury W., Gutierrez C., et al. Molecular characteristics and spatial distribution of adult human corneal cell subtypes // Sci Rep. 2021. Vol. 11, N 1. P. 16323. doi: 10.1038/s41598-021-94933-8

[9] Campos M, Szerenyi K, Lee M, et al. Keratocyte loss after corneal deepithelialization in primates and rabbits. Arch Ophthalmol. 1994;112(2):254. doi: 10.1001/archopht.1994.01090140130034

[10] Campos M., Szerenyi K., Lee M., et al. Keratocyte loss after corneal deepithelialization in primates and rabbits // Arch Ophthalmol. 1994. Vol. 112, N 2. P. 254. doi: 10.1001/archopht.1994.01090140130034

[11] Wilson S, He Y, Weng J, et al. Epithelial injury induces keratocyte apoptosis: hypothesized role for the interleukin-1 system in the modulation of corneal tissue organization and wound healing. Exp Eye Res. 1996;62(4):325–338. doi: 10.1006/exer.1996.0038

[12] Wilson S., He Y., Weng J., et al. Epithelial injury induces keratocyte apoptosis: hypothesized role for the interleukin-1 system in the modulation of corneal tissue organization and wound healing // Exp Eye Res. 1996. Vol. 62, N 4. P. 325–338. doi: 10.1006/exer.1996.0038

[13] Zieske J, Guimarães, S, Hutcheon A. Kinetics of keratocyte proliferation in response to epithelial debridement. Exp Eye Res. 2001;72(1):33–39. doi: 10.1006/exer.2000.0926

[14] Zieske J., Guimarães S., Hutcheon A. Kinetics of keratocyte proliferation in response to epithelial debridement // Exp Eye Res. 2001. Vol. 72, N 1. P. 33–39. doi: 10.1006/exer.2000.0926

[15] Fini ME, Stramer BM. How the cornea heals: cornea-specific repair mechanisms affecting surgical outcomes. Cornea. 2005;24(8 Suppl):S2–S11. doi: 10.1097/01.ico.0000178743.06340.2c

[16] Fini M.E., Stramer B.M. How the cornea heals: cornea-specific repair mechanisms affecting surgical outcomes // Cornea. 2005. Vol. 24, N 8 Suppl. P. S2–S11. doi: 10.1097/01.ico.0000178743.06340.2c

[17] Klenkler B, Sheardown H, Jones L. Growth factors in the tear film: role in tissue maintenance, wound healing, and ocular pathology. Ocul Surf. 2007;5(3):228–239. doi: 10.1016/s1542-0124(12)70613-4

[18] Klenkler B., Sheardown H., Jones L. Growth factors in the tear film: role in tissue maintenance, wound healing, and ocular pathology // Ocul Surf. 2007. Vol. 5, N 3. P. 228–239. doi: 10.1016/s1542-0124(12)70613-4

[19] Anitua E, de la Sen-Corcuera B, Orive G, et al. Progress in the use of plasma rich in growth factors in ophthalmology: from ocular surface to ocular fundus. Expert Opin Biol Ther. 2022;22(1):31–45. doi: 10.1080/14712598.2021.1945030

[20] Anitua E., de la Sen-Corcuera B., Orive G., et al. Progress in the use of plasma rich in growth factors in ophthalmology: from ocular surface to ocular fundus // Expert Opin Biol Ther. 2022. Vol. 22, N 1. P. 31–45. doi: 10.1080/14712598.2021.1945030

[21] Anitua E, Alonso R, Girbau C, et al. Antibacterial effect of plasma rich in growth factors (PRGF®-Endoret®) against Staphylococcus aureus and Staphylococcus epidermidis strains. Clin Exp Dermatol. 2012;37(6):652–657. doi: 10.1111/j.1365-2230.2011.04303.x

[22] Anitua E., Alonso R., Girbau C., et al. Antibacterial effect of plasma rich in growth factors (PRGF®-Endoret®) against Staphylococcus aureus and Staphylococcus epidermidis strains // Clin Exp Dermatol. 2012. Vol. 37, N 6. P. 652–657. doi: 10.1111/j.1365-2230.2011.04303.x

[23] Anitua E, Muruzabal F, de la Fuente M, et al. PRGF exerts more potent proliferative and anti-inflammatory effects than autologous serum on a cell culture inflammatory model. Exp Eye Res. 2016;151:115–121. doi: 10.1016/j.exer.2016.08.012

[24] Anitua E., Muruzabal F., de la Fuente M., et al. PRGF exerts more potent proliferative and anti-inflammatory effects than autologous serum on a cell culture inflammatory model // Exp Eye Res. 2016. Vol. 151. P. 115–121. doi: 10.1016/j.exer.2016.08.012

[25] Sanchez-Avila RM, Merayo-Lloves J, Riestra AC, et al. The effect of immunologically safe plasma rich in growth factor eye drops in patients with sjögren syndrome. J Ocul Pharmacol Ther. 2017;33(5):391–399. doi: 10.1089/jop.2016.0166

[26] Sanchez-Avila R.M., Merayo-Lloves J., Riestra A.C., et al. The effect of immunologically safe plasma rich in growth factor eye drops in patients with Sjögren syndrome // J Ocul Pharmacol Ther. 2017. Vol. 33, N 5. P. 391–399. doi: 10.1089/jop.2016.0166

[27] Foster TE, Puskas BL, Mandelbaum BR, et al. Platelet-rich plasma: from basic science to clinical applications. Am J Sports Med. 2009;37(11):2259–2272. doi: 10.1177/0363546509349921

[28] Foster T.E., Puskas B.L., Mandelbaum B.R., et al. Platelet-rich plasma: from basic science to clinical applications // Am J Sports Med. 2009. Vol. 37, N 11. P. 2259–2272. doi: 10.1177/0363546509349921

[29] Park YB, Kim JH, Ha CW, Lee DH. Clinical efficacy of platelet-rich plasma injection and its association with growth factors in the treatment of mild to moderate knee osteoarthritis: a randomized double-blind controlled clinical trial as compared with hyaluronic acid. Am J Sports Med. 2021;49(2):487–496. doi: 10.1177/0363546520986867

[30] Park Y.B., Kim J.H., Ha C.W., Lee D.H. Clinical efficacy of platelet-rich plasma injection and its association with growth factors in the treatment of mild to moderate knee osteoarthritis: a randomized double-blind controlled clinical trial as compared with hyaluronic acid // Am J Sports Med. 2021. Vol. 49, N 2. P. 487–496. doi: 10.1177/0363546520986867

[31] Wilson SE. Corneal wound healing. Exp Eye Res. 2020;197:108089. doi: 10.1016/j.exer.2020.108089

[32] Wilson S.E. Corneal wound healing // Exp Eye Res. 2020. Vol. 197. P. 108089. doi: 10.1016/j.exer.2020.108089

[33] Kasparov AA, Kasparova EvgA, Fadeeva LL, et al. Personalized cell therapy for early postoperative bullous keratopathy (experimental proof and clinical results). Vestnik Oftalmologii. 2013;129(5):53–61. (In Russ).

[34] Каспаров А.А., Каспарова Евг.А., Фадеева Л.Л., и др. Персонализированная клеточная терапия ранней буллезной кератопатии (экспериментальное обоснование и клинические результаты) // Вестник офтальмологии. 2013. Т.129, № 5. С. 53–61.

[35] Koval’chuk LV, Pavlyuk AS, Kasparov AA, et al. Analysis of pharmacological agents in the model of apoptosis of human lymphocytes in vitro in normal conditions and in immunopathology. Allergology and Immunology. 2000;1:24–30. (In Russ).

[36] Ковальчук Л.В., Павлюк А.С., Каспаров А.А., и др. Анализ фармакологических средств на модели апоптоза лимфоцитов человека in vitro в норме и при иммунопатологии // Аллергология и иммунология. 2000. № 1. С. 24–30.

[37] Seidelmann N, Duarte Campos DF, Rohde M, et al. Human platelet lysate as a replacement for fetal bovine serum in human corneal stromal keratocyte and fibroblast culture. J Cell Mol Med. 2021;25(20):9647–9659. doi: 10.1111/jcmm.16912

[38] Seidelmann N., Duarte Campos D.F., Rohde M., et al. Human platelet lysate as a replacement for fetal bovine serum in human corneal stromal keratocyte and fibroblast culture // J Cell Mol Med. 2021. Vol. 25, N 20. P. 9647–9659. doi: 10.1111/jcmm.16912

[39] Anitua E, de la Fuente M, Muruzabal F, et al. Plasma rich in growth factors (PRGF) eye drops stimulates scarless regeneration compared to autologous serum in the ocular surface stromal fibroblasts. Exp Eye Res. 2015;135:118–126. doi: 10.1016/j.exer.2015.02.016

[40] Anitua E., de la Fuente M., Muruzabal F., et al. Plasma rich in growth factors (PRGF) eye drops stimulates scarless regeneration compared to autologous serum in the ocular surface stromal fibroblasts // Exp Eye Res. 2015. Vol. 135. P. 118–126. doi: 10.1016/j.exer.2015.02.016

[41] Koulikovska M, Szymanowski O, Lagali N, Fagerholm P. Platelet-rich plasma prolongs myofibroblast accumulation in corneal stroma with incisional wound. Curr Eye Res. 2015;40(11):1102–1110. doi: 10.3109/02713683.2014.978478

[42] Koulikovska M., Szymanowski O., Lagali N., Fagerholm P. Platelet-rich plasma prolongs myofibroblast accumulation in corneal stroma with incisional wound // Curr Eye Res. 2015. Vol. 40, N 11. P. 1102–1110. doi: 10.3109/02713683.2014.978478

RIGHTS & PERMISSIONS

Eco-Vector