Percutaneous application of allogeneic adipose-derived mesenchymal stem cell in dogs submitted to minimally invasive plate osteosynthesis of the tibia

Franco, Guilherme Galhardo; Minto, Bruno Watanabe; Dreibi, Rafael Manzini; Costa Junior, Jose Sergio; Dias, Luis Gustavo Gosuen Gonçalves

doi:10.1590/ACB360206

person Guilherme Galhardo Franco

schoolDVM, MSc, PhD. Universidade Federal do Espírito Santo – Center for Agricultural Sciences and Engineering – Department of Veterinary Medicine – Alegre (ES), Brazil.

0000-0001-9945-3453

person Bruno Watanabe Minto *

schoolDVM, MSc, PhD. Universidade Estadual Paulista – Faculty of Agrarian and Veterinary Sciences – Department of Veterinary Clinic and Surgery – Jaboticabal (SP), Brazil.

0000-0002-8808-4127

person Rafael Manzini Dreibi

schoolDVM, MSc. Universidade Estadual Paulista – Faculty of Agrarian and Veterinary Sciences – Department of Veterinary Clinic and Surgery – Jaboticabal (SP), Brazil.

0000-0002-7705-6075

person Jose Sergio Costa Junior

schoolDVM, MSc. Universidade Estadual Paulista – Faculty of Agrarian and Veterinary Sciences – Department of Veterinary Clinic and Surgery – Jaboticabal (SP), Brazil.

0000-0002-4725-5962

person Luis Gustavo Gosuen Gonçalves Dias

schoolDVM, MSc, PhD. Universidade Estadual Paulista – Faculty of Agrarian and Veterinary Sciences – Department of Veterinary Clinic and Surgery – Jaboticabal (SP), Brazil.

0000-0003-3993-1649

person*Corresponding author:
brunowminto@gmail.com | (55 16)3209-7100

Conflict of interest: Nothing to declare.

groupsAuthors’ contribution

Design the study: Franco GG, Minto BW, Dreibi RM, Costa Junior JS and Dias LGGG; Critical revision: Franco GG, Minto BW, Dias LGGG and Costa Junior JS; Technical procedures: Franco GG and Minto BW; Acquisition of data: Franco GG and Dreibi RM; Final approval: Franco GG and Minto BW.

Figures (4)
Tables (2)

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Figure 1 Intraoperative photographs of the minimally invasive osteosynthesis procedure with plate and application of mesenchymal stem cells derived from adipose tissue (a–d) and radiographic sequence in the craniocaudal projection (e–i) of a dog (dog 20), without defined race (MBD), 15 years of age, with a body mass of 13 kg and distal metaphyseal fracture of tibia in spiral, complete, closed with the fractured fibula. The fracture was indirectly reduced by manual traction (a) and the plate was inserted through the proximal access and slipped through the epiperiosteal tunnel over the medial surface of the tibia (b) and the bone perforations for application of the screws were performed by distal and proximal accesses. A needle was inserted into the joint space to delimit the tibia (c). After dermorrhaphy, 3 million allogeneic AD-MSC diluted in 2 mL of 0.9% saline solution were percutaneously applied to the focus of the fracture using a hypodermic needle and syringe (d). The distal metaphyseal spiral tibial fracture (e) was treated with a 2.7 mm 12-hole locked plate, applying three proximal locked screws and two distal locked screws (f). Radiographic follow-up of the fracture at 15 (g), 30 (h) and 60 days (i), demonstrating bone healing.

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Figure 2 Pictures of the minimally invasive osteosynthesis procedure with plate and application of mesenchymal stem cells derived from adipose tissue (a–d) and radiographic images (e-h) of a 5-year-old dog without a defined race, body mass of 7.1 kg with spiral diaphyseal fracture of the tibia in the middle third with the fractured fibula. The fracture was indirectly reduced with the aid of bone tweezers applied to the bone through the proximal and distal approaches (a). MSC-AD washed in 5 mL of cell wash solution using a pipette (b). Photograph showing the cellular button (orange arrow) formed after centrifugation at 1250 RPM for 3 min performed in the process of thawing and cell washing (c). After dermorrhaphy, 3 million allogeneic AD-MSC diluted in 2 mL of 0.9% saline solution were percutaneously applied to the focus of the fracture using a hypodermic needle and syringe (d). Preoperative radiographic images of the fracture (e, f) and in the immediate postoperative period, demonstrating the correction of valgus and recurvatum deviations (g, h).

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Figure 3 Craniocaudal and mediolateral radiographic images of the tibia of the dog G1-14, without defined race of 2 years of age, with body mass 23.2 kg proximal metaphyseal fracture of the tibia, comminuted and with the fibula fractured in the preoperative period (a, b). The immediate postoperative radiographs (c, d) demonstrated the complication of screw insertion in the knee joint space (orange arrow). Craniocaudal radiographic projections with 60 days (e) and 90 days (f) postoperatively, demonstrating good bone healing. The most proximal screw is observed without penetrating the joint space after replacement (e, f).

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Figure 4 Craniocaudal radiographic images of the tibia of the dog 11 from the control group (mongrel, 5 years of age and 23 kg). Note a mid-diaphyseal comminuted fracture of tibia and fibula in the immediate postoperative period (a). Craniocaudal radiographic projections after 15 (b), 30 (c), 60 (d) and 90 days postoperatively (e). Note the radiographic clinical union at 120 days postoperatively (f).

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Table 1 Summary of the clinical data of the dogs in both groups included in this study with tibial fractures treated with MIPO and in the G2 percutaneous injection of the allogeneic AD-MSC.

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Table 2 Median ± interquartile range (IQR) of dogs from group 1 (percutaneous application of allogeneic AD-MSC at the fracture site) and group 2 (control).

Dogs	Age (months)	Breed	Weight (kg)	Fracture	Implants	Bone union (days)
G1-1	3	Dober.	5.6	Diaf., trans., middle	2.0 mm, 8 H., 4 screws	15
G1-2	3	MBD	24.2	Diaf., obliq., distal	3.5 mm, 11 H., 4 screws	30
G1-3	5	Boxer	10	Diaf., obliq., middle	2.0 mm, 10 H., 4 screws	15
G1-4	7	MBD	6.3	Diaf., obliq., middle	2.0 mm, 10 H., 4 screws	15
G1-5	7	MBD	7.1	Diaf., obliq., middle	2.0 mm, 9 H., 4 screws	15
G1-6	8	Shih Tzu	6.6	Diaf., spiral, middle	2.0 mm, 8 H., 4 screws	30
G1-7	9	MBD	7.9	Diaf., trans., middle	2.0 mm, 10 H., 4 screws	60
G1-8	9	MBD	20.5	Diaf., spiral, middle	3.5 mm, 11 H., 4 screws	15
G1-9	9	Eng. Bull.	16	Diaf., spiral, middle	2.7 mm, 9 H., 5 screws	30
G1-10	11	Am.Staf.Ter.	27	Diaf., obliq., prox.	3.5 mm, 10 H., 4 screws	30
G1-11	12	Rott.	25.8	Diaf., obliq., middle	2.7 mm, 12 H., 5 screws	15
G1-12	12	Bull Ter.	14	Diaf., obliq., middle	2.7 mm, 11 H., 5 screws	30
G1-13	24	MBD	17	Diaf., spiral, middle	2.7 mm, 8 H., 4 screws	30
G1-14	24	MBD	23.2	Meta., com., prox.	3.5 mm, 12 H., 5 screws	30
G1-15	48	C.Chow	25.2	Diaf., com., middle	3.5 mm, 11 H., 5 screws	30
G1-16	60	MBD	7.1	Diaf., spiral, middle	2.0 mm, 9 H., 5 screws.	30
G1-17	96	MBD	18	Diaf., com., middle	2.7 mm, 12 H., 5 screws	30
G1-18	156	Rott.	36	Meta., com., distal	3.5 mm B, 14H., 6 screws	30
G1-19	156	Pinscher	2.7	Diaf., obliq., prox.	1.5 mm, 10 H., 4 screws	60
G1-20	180	MBD	13	Meta., spiral., distal	2.7 mm, 12 H., 5 screws	30
G2-1	4	Poodle	5.2	Diaf., trans., middle	2.0 mm, 10H., 3 screws	60
G2-2	7	MBD	10	Diaf., trans., middle	2.7 mm, 7H., 2 screws	90
G2-3	8	Border Collie	25	Diaf., obliq., middle	3.5 mm, 10H., 2screws	60
G2-4	8	Labrador Retriever	20	Diaf., obliq., middle	3.5 mm, 11H., 2 screws	15
G2-5	8	Labrador Retriever	30	Diaf., trans., prox.	3.5 mm, 11H., 3 screws	60
G2-6	9	MBD	16	Diaf., com., middle	2.7 mm, 10H., 3 screws	30
G2-7	11	Blue Heeler	15	Diaf., trans., middle	2.7 mm, 10H., 3 screws	60
G2-8	12	MBD	20	Diaf. com., middle	2.7 mm, 9H., 2 screws	30
G2-9	12	MBD	30	Diaf., trans., middle	3.5 mm, 11H., 3 screws	60
G2-10	48	MBD	8	Diaf., com., middle	2.7 mm, 7H., 2 screws	90
G2-11	60	MBD	23	Diaf., com., middle	3.5 mm, 12H., 2 screws	120
G2-12	60	MBD	20	Diaf., trans., middle	4.5 mm, 12H., 3 screws	90
G2-13	60	MBD	5	Diaf., trans., prox.	2.0 mm, 10H., 3 screws	90
G2-14	72	Dachshund	7	Diaf., trans., middle	2.0 mm, 7H., 2 screws	120
G2-15	108	Dachshund	9	Diaf., trans., middle	2.4 mm, 7H., 2 screws	60
G2-16	120	MBD	5.5	Diaf., trans., distal	2.0 mm, 9H., 2 screws	90

	Group 1 (n =20)		Group 2 (n =16)		P-value
	Median	IQR	Median	IQR	P-value
Age, months	11.5	43.25	12.0	52.00	0.7015
Weight, kg	15.0	16.35	15.5	13.00	0.9112
Bone union, days	30.0b	15.00	60.0a	30.00	< 0.0001

[1] G1. Group 1 – mesenchymal stem cell group; G2. Group 2 – Control group; y., years; m., months; MBD, mixed breed dog; Rott., Rottweiler; Bull Ter., Bull Terrier; Dober., Dobermann Pinscher.; Am.Staf.Ter., American Staffordshire Terrier; C.Chow, Chow-Chow; Eng. Bull., English Bulldog; diaf., diaphyseal; meta., metaphyseal; obliq., oblique; trans., transverse; com., comminuted; B., Broad; H., holes; kg, kilogram.