Dynamics of the number of myocyte nuclei in muscle portions of arterial and venous homograft walls during long-term preservation

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Abstract

INTRODUCTION: An important issue of use of freshly prepared homograft’s in the reconstructive vascular surgery is the maximal length of their preservation for safe application. Histological examinations are required.

AIM: To determine the optimal timing of the use of homograft’s based on the dynamics of the number of myocyte nuclei of muscle portions of arterial and venous homograft walls preserved in RPMI 1640 solution with the addition of gentamicin and fluconazole at a temperature of +4°C.

MATERIALS AND METHODS: The study of arterial and venous homograft’s from a posthumous donor was performed in compliance with the rules for collecting organs and tissues. The homograft’s were preserved in RPMI 1640 solution with the addition of gentamicin (400 μg/ml) and fluconazole (20 μg/ml) at a temperature of +4°C. At intervals of 7 days up to 84 days, sections of the arterial and venous grafts were cut out and fixed in formalin. Then glass slides were prepared. A total of 120 arterial and 120 venous samples were studied. The number of myocyte nuclei in 0.01 mm2 muscle portions of the walls was estimated.

RESULTS: On day 42, a decrease in the number of myocyte nuclei in arterial homograft’s by 56% was recorded relative to the values of day 7 (p = 0.003), a decrease in the number of myocyte nuclei in venous homograft’s by 55% (p = 0.024); a decrease in nuclei was also recorded at all subsequent control points.

CONCLUSION: The optimal period for the use of arterial homograft’s can be considered the period of preservation in RPMI 1640 solution for up to 21 days. Venous homograft’s can be used for up to 42 days, despite the decrease in the number of myocyte nuclei after 35 days, since there is no edema of the venous homograft wall until the end of the 42 day.

About the authors

Roman E. Kalinin

Ryazan State Medical University

Email: kalinin-re@yandex.ru
ORCID iD: 0000-0002-0817-9573
SPIN-code: 5009-2318

MD, Dr. Sci. (Medicine); Professor

Russian Federation, Ryazan

Igor A. Suchkov

Ryazan State Medical University

Email: suchkov_med@mail.ru
ORCID iD: 0000-0002-1292-5452
SPIN-code: 6473-8662

MD, Dr. Sci. (Medicine), Professor

Russian Federation, Ryazan

Vyacheslav V. Karpov

Ryazan State Medical University; Regional Clinical Hospital

Author for correspondence.
Email: sdrr.s@yandex.ru
ORCID iD: 0000-0001-5523-112X
SPIN-code: 6245-6292

MD, Cand. Sci. (Medicine)

Russian Federation, Ryazan; Ryazan

Alexander P. Shvalb

Regional Clinical Tuberculosis Dispensary

Email: shvalbalik@yandex.ru
ORCID iD: 0000-0001-8563-0439

MD, Cand. Sci. (Medicine)

Russian Federation, Ryazan

Dmitry V. Guzairov

Ryazan State Medical University

Email: sdrr@mail.ru
ORCID iD: 0009-0002-1943-2244
SPIN-code: 3501-8157
Russian Federation, Ryazan

Tatyana M. Cherdantseva

Ryazan State Medical University

Email: cherdan.morf@yandex.ru
ORCID iD: 0000-0002-7292-4996
SPIN-code: 3773-8785

MD, Dr. Sci. (Medicine), Professor

Russian Federation, Ryazan

References

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  2. Kalinin RE, Suchkov IA, Karpov VV, et al. The First Experience of Using Allografts in Surgery of Main Arteries of Lower Limbs. Russian Sklifosovsky Journal “Emergency Medical Care”. 2020;9(3):442–448. doi: 10.23934/2223-9022-2020-9-3-442-448 EDN: IZFWQX
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Supplementary files

Supplementary Files
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1. JATS XML
2. Fig. 1. The stages of digital processing of the homograph wall (magnified ×100).

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3. Fig. 4. Fragmentation and lysis of smooth myocyte nuclei in a homograph. Indicated by an arrow (magnified × 400).

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4. Fig. 5. Arterial homograph, day 7 of preservation in RPMI 1640 solution. Hematoxylin-eosin staining, digital magnification: endotheliocytes are partially preserved, their nuclei are hyperchromic, swollen, moderate edema of the intima, the inner elastic membrane is well differentiated, the cleavage sites of the inner elastic membrane, the contours of leiomyocytes are clearly defined, the nuclei are not swollen, vacuolization of individual myocytes (voids around the nuclei in the middle layer).

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5. Fig. 6. Arterial homograph, day 7 of preservation in RPMI 1640 solution. Weigert–Van Gieson coloring, digital magnification: dark blue, well-defined inner elastic membrane in the form of two monolines, wavy thin elastic fibers in the middle layer, collagen in the form of a pinkish background without fiber visualization.

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6. Fig. 7. Venous homograph, 7 days of preservation in RPMI 1640 solution. Hematoxylin-eosin staining, digital magnification: the endothelium is well preserved, the intima is wide, with thick collagen fibers, the myocytes of the middle layer are clearly differentiated; the well-preserved endothelium vasa vasorum attracts attention.

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7. Fig. 8. Venous homograph, 7 days of preservation in RPMI 1640 solution. Weigert–Van Gieson coloring, digital magnification: collagen is well expressed in the form of thick fibers of purple shades.

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8. Fig. 9. Arterial homograph, 21 days of preservation in RPMI 1640 solution. Hematoxylin-eosin staining, digital magnification: endotheliocytes are preserved in places, the basement membrane is clearly differentiated, the intima is thin, the inner elastic membrane is in the form of a wide red split line, the contours of smooth myocytes are preserved in places, the nuclei are thin, arranged with uneven density, forming areas of devastation.

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9. Fig. 10. Arterial homograph, 21 days of preservation in RPMI 1640 solution. Weigert–Van Gieson coloring, digital magnification: the split inner elastic membrane is clearly defined, thin blue (elastic) filaments in the middle shell, with faintly noticeable eosinophilic collagen between them.

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10. Fig. 11. Venous homograph, 21 days of preservation in RPMI 1640 solution. Hematoxylin-eosin staining, digital magnification: endotheliocytes are preserved in places, their nuclei are hyperchromic, the basement membrane is well differentiated, the intima is wide and dense, the contours of the myocytes are predominantly preserved, the endothelium vasa vasorum is preserved.

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11. Fig. 12. Venous homograph, 21 days of preservation in RPMI 1640 solution. Weigert–Van Gieson coloring, digital magnification: collagen is pale colored in intimacy and well perceives the dye in the middle shell.

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12. Fig. 13. Arterial homograph, 84 days of preservation in RPMI 1640 solution. Hematoxylin-eosin staining, digital magnification: single endotheliocytes with a flat pale nucleus, the basement membrane is fragmented, with areas of detachment, the intima is compacted, the inner elastic membrane in the form of fragments of different lengths and thicknesses; single myocytes, some retain contours, pale nuclei, wide areas of devastation.

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13. Fig. 14. Arterial homograph, 84 days of preservation in RPMI 1640 solution. Weigert–Van Gieson coloring, digital magnification: intima fibers are pale, disconnected, the inner elastic membrane is in the form of a clear monoline, in the middle shell there is a light purple fibrous background of different coloring densities.

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14. Fig. 15. Venous homograph, 84 days of preservation in RPMI 1640 solution. Hematoxylin-eosin staining, digital magnification: endotheliocytes are preserved in many areas, the basement membrane is thin, the contours of the myocytes of the middle shell are predominantly preserved, the nuclei are pale, not everywhere, multiple small areas of devastation, the endothelium vasa vasorum is not determined.

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15. Fig. 16. Venous homograph, 84 days of preservation in RPMI 1640 solution. Weigert–Van Gieson coloring, digital magnification: pale coloration of collagen fibers in light red shades in the intima and muscle layer.

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16. Fig. 2. Dynamics of the number of myocyte nuclei in 0.01 mm2 arterial graft (M±SD).

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17. Fig. 3. Dynamics of the number of myocyte nuclei in 0.01 mm2 of venous graft (M±SD).

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