Morphological evaluation of decellularized lyophilized amniotic membrane

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Abstract

Aim – to study the morphological structure of lyophilized amniotic membrane preliminarily subjected to physical decellularization.

Material and methods. An experimental study of the preservation of the anatomical structure of lyophilized amniotic membrane was performed on four groups of amniotic membrane fragments. Group 1: AM impregnated with glycerin and dried over silica gel; Group 2: AM impregnated with glycerin, treated ultrasonically and lyophilized; Group 3: AM treated ultrasonically and lyophilized; Group 4: native AM without preservation. The biomaterial was studied using light microscopy and scanning electron microscopy.

Results. Physical methods of influencing biological tissue have an expected effect on cell viability and allow obtaining a completely decellularized amniotic membrane. Additional treatment with glycerol before physical action on biological tissue for the purpose of decellularization does not have a significant effect on the preservation of cellular structures. It should only be noted that in the amniotic membrane impregnated with glycerol, more fragments of epithelial cell membranes are preserved and the basement membrane is more preserved.

Conclusion. The decellularization method developed by us using physical methods does not introduce any chemicals into the processed biomaterial that can have an unpredictable effect on regenerating tissues. Preservation of the amniotic membrane by lyophilization allows obtaining a morphologically integral, elastic and durable biomaterial.

About the authors

Kseniya E. Kuchuk

Samara Regional Clinical Ophthalmological Hospital named after T.I. Eroshevsky

Email: kuchukke@rambler.ru
ORCID iD: 0009-0003-2986-5913

MD, ophthalmologist, head of the tissue procurement and preservation department

Russian Federation, Samara

Larisa T. Volova

Samara State Medical University

Email: l.t.volova@samsmu.ru
ORCID iD: 0000-0002-8510-3118

MD, Dr. Sci. (Medicine), Professor, Director of the “BioTech” Research Institute

Russian Federation, Samara

Iosif V. Novikov

Samara State Medical University

Email: р111аа@yandex.ru
ORCID iD: 0000-0002-6855-6828

MD, Cand. Sci. (Medicine), assistant of the Department of Traumatology, Orthopedics and Extreme Surgery named after Academician of the Russian Academy of Sciences A.F. Krasnov

Russian Federation, Samara

Evgenii S. Milyudin

Samara State Medical University

Author for correspondence.
Email: e.s.milyudin@samsmu.ru
ORCID iD: 0000-0001-7610-7523

MD, Dr. Sci. (Medicine), Associate professor, Department of Operative Surgery and Clinical Anatomy with a course in Medical Information Technologies

Russian Federation, Samara

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2. Figure 1. Native amniotic membrane preparation. Stained with picrofuchsin. Magnification x400.

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3. Figure 2. Amniotic membrane preparation preserved by drying over silica gel after preliminary treatment with glycerol. Stained with hematoxylin and eosin. Magnification x400.

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4. Figure 3. Electron microscopic image of the amniotic membrane in a scanning electron microscope. Epithelial surface of the amniotic membrane preparation dried with silica and pre-impregnated with glycerol. Magnification x1000.

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5. Figure 4. Electron microscopic image of the amniotic membrane in a scanning electron microscope. Spongy layer of the amniotic membrane preparation dried with silica and preliminary impregnation with glycerin. Magnification x400.

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6. Figure 5. Amniotic membrane preparation preserved by lyophilization after preliminary treatment with glycerol. Stained with hematoxylin and eosin. Magnification x400.

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7. Figure 6. Electron microscopic image of amniotic membrane in a scanning electron microscope. Epithelial surface of a lyophilized amniotic membrane preparation with preliminary impregnation with glycerol. Magnification x50.

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8. Figure 7. Electron microscopic image of amniotic membrane in a scanning electron microscope. Spongy layer of a lyophilized amniotic membrane preparation with preliminary impregnation with glycerol. Magnification x400.

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9. Figure 8. Amniotic membrane preparation preserved by lyophilization without glycerol treatment. Hematoxylin and eosin staining. Magnification x400.

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10. Figure 9. Electron microscopic image of amniotic membrane in a scanning electron microscope. Epithelial surface of a lyophilized amniotic membrane preparation without preliminary impregnation with glycerol. Magnification x400.

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11. Figure 10. Electron microscopic image of amniotic membrane in a scanning electron microscope. Spongy layer of the preparation of lyophilized amniotic membrane without preliminary impregnation with glycerol. Magnification x1000.

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Copyright (c) 2025 Kuchuk K.E., Volova L.T., Novikov I.V., Milyudin E.S.

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