Center for Regenerative Medicine Research,
Saga University School of Medicine

佐賀大学医学部附属 再生医学研究センター

Scaffold-Free
Biofabrication
of Living Organs

Pioneering the Kenzan method — a robotic bioprinting technology that assembles living tissue constructs entirely from cells, without any artificial scaffolds. Saga University School of Medicine.

The Kenzan method: cells form spheroids, which are placed onto a microneedle array, fuse into continuous tissue, and are harvested as a scaffold-free construct
Note: There is another Koichi Nakayama at Saga University in Information Engineering. Please ensure you are contacting the correct laboratory.
*同姓同名に注意！
佐賀大学には同姓同名の中山功一教授（佐賀大学大学院 工学系研究科 知能情報システム学専攻）が在籍していますので書類等送付時は所属など宛先をご確認ください。
The Kenzan Method

How It Works

01

Spheroid Formation

Cells are cultured into uniform multicellular spheroids — the fundamental building blocks of our constructs.

02

Robotic Placement

A Bio-3D printer precisely skewers spheroids onto a microneedle array (Kenzan) according to a 3D design.

03

Fusion & Maturation

Spheroids fuse spontaneously through natural cell adhesion, forming a continuous living tissue.

04

Construct Harvest

The fused construct is removed from the needle array — entirely scaffold-free, made only from living cells.

Research Output

Publications

2026
Biofabrication strategies for engineered constructs in cardiovascular tissue repair
Yu J, Murata D, Itoh M, Nakayama K
Regenerative Therapy 32, 101110
2026
Extracellular Matrix Microspheres with Magnetically Labelled MSCs enable Functional Regeneration of the Osteo-Tendinous Junction
Datla A, Bairagya G, Nakayama K, Vadrevu S, Rath SN
ACS Appl. Mater. Interfaces 2026
2026
Modulation of iPSC-derived NCMSC cell state by TD-198946 enhances scaffold-free cartilage biofabrication
Nakamura A, Zujur D, Nonaka T, Yoshizato H, Kashimoto S, Ikeya M, Nakayama K
Biofabrication 18 025004
2026
A Scaffold-Free Cartilage Construct Fabricated Using a Bio 3D Printer Accelerates Critical-Size Bone Defect Regeneration
Yoshizato H, Murata D, Kashimoto S, Nonaka T, Fujimoto R, Nagaishi Y, Itoh M, Morimoto T, Nakayama K
J Orthopaedic Translation 57, 101033
2025
Response to the letter to the editor regarding "Bio-3D printing of scaffold-free ADSC-derived cartilage constructs comparable to natural cartilage in vitro"
Nonaka T, Murata D, Yoshizato H, Kashimoto S, Nakamura A, Morimoto T, Nakayama K
J Orthop Surg Res 2025
2025
Bio-3D printing with smooth muscle cells derived from human iPSCs via neural crest and its application for the tracheal regeneration
Hashimoto S, Taniguchi D, Doi R, Obata T, Shiraishi T, Matsumoto T, et al.
Biofabrication 18(1)
2025
Efficacy and Safety of Bio 3D Conduits Composed of Human Umbilical Cord–Derived Mesenchymal Stromal Cells: A Proof-of-Concept Study in a Canine Ulnar Nerve Defect Model
Fujita K, Ikeguchi R, Aoyama T, Noguchi T, Yoshimoto K, Sakamoto D, et al.
Cell Transplantation 34
2025
Artificial Blood Vessels — Clinical Development of TEVG Review
Itoh M, Kamohara K, Node K, Nakayama K
J Artificial Organs 2025
2025
Xenograft of bio-3D printed scaffold-free cartilage constructs derived from human iPSCs to regenerate articular cartilage in immunodeficient pigs
Nonaka T, Murata D, Nakamura A, Yoshizato H, Kashimoto S, Nagaishi Y, Itoh M, Zujur D, Ikeya M, Toguchida J, Mawatari M, Nakayama K
Regenerative Therapy 29, 506–516
2025
Promotion of bone-tendon healing after ACL reconstruction using scaffold-free constructs comprising ADSCs produced by a bio-3D printer in rabbit models
Higa K, Murata D, Azuma C, Nishida K, Nakayama K
J Orthopaedic Translation 52, 265–275
2025
Bio-3D printing of scaffold-free ADSC-derived cartilage constructs comparable to natural cartilage in vitro
Nonaka T, Murata D, Yoshizato H, Kashimoto S, Nakamura A, Morimoto T, Nakayama K
J Orthop Surg Res 20, 182
2024
Fabrication of a Bio-3D Printed Scaffold-Free Artificial Trachea with Horseshoe-Shaped Hyaline Cartilage
Uchida F, Matsumoto K, Nishimuta M, et al.
European J Cardio-Thoracic Surgery 66(4)
2024
Nerve regeneration using a Bio 3D conduit derived from umbilical cord-derived mesenchymal stem cells
Iwai T, Aoyama T, Noguchi T, et al.
PLoS ONE 19(12):e0310711
2024
Creating 3D constructs with cranial neural crest-derived cell lines using a bio-3D printer
Taguchi M, Yoshimoto S, Suyama K, et al.
J Oral Biosciences 66(2), 339–348
2024
Three-Dimensional Bio-Printed Tubular Tissue Using Dermal Fibroblast Cells as a New Tissue-Engineered Vascular Graft
Hayasaka M, Kokudo T, Kaneko J, et al.
ASAIO Journal 70(11), 1008–1014
2024
Scaffold-free bone-like 3D structure established through osteogenic differentiation from human gingiva-derived stem cells
Toyoda M, Fukuda T, Fujimoto R, et al.
Biochemistry and Biophysics Reports 38, 101656
2023
Special Issue: Biofabrication with Spheroid and Organoid Materials Guest Editor
Skylar-Scott M, Declercq H, Nakayama K
Acta Biomaterialia 165
2023
Scaffold-free human vascular calcification model using a bio-three-dimensional printer
Nagaishi Y, Murata D, Yoshizato H, Nonaka T, Itoh M, Hara H, Nakayama K
Biofabrication 15(4)
2023
Enhanced chondrogenic differentiation of iPS cell-derived mesenchymal stem/stromal cells via neural crest cell induction
Zujur D, Al-Akashi Z, Nakamura A, et al.
Frontiers in Cell and Developmental Biology 11:1140717
2023
Regeneration of the ureter using a scaffold-free live-cell structure created with bio-three-dimensional printing
Takagi K, Taniguchi D, Machino R, et al.
Acta Biomaterialia 165, 102–110
2022
3D Printing in Nephrology Chapter
Nonaka T, Nagaishi Y, Murata D, Hara H, Nakayama K
In: Innovations in Nephrology, Springer
2022
Involvement of the PI3K/AKT Pathway in the Formation and Fusion of Spheroids Derived from Human Dermal Fibroblast for Tissue Engineering Technology
Amamoto S, Itoh M, Takahashi B, Kitsuka T, Uchiashi K, Murata D, Node K, Nakayama K, Kamohara K
Cell and Tissue Biology 16, 312–329
2022
Nerve regeneration using the Bio 3D conduit fabricated with spheroids Review
Ikeguchi R, Aoyama T, Noguchi T, et al.
J Artificial Organs 25, 289–297
2022
Bio-3D printing of scaffold-free osteogenic and chondrogenic constructs using rat adipose-derived stromal cells
Fujimoto R, Murata D, Nakayama K
Front. Biosci. (Landmark Ed) 27(2), 052
2022
Scaffold-Free Tubular Engineered Heart Tissue From Human iPSCs Using Bio-3D Printing Technology In Vivo
Kawai Y, Tohyama S, Arai K, et al.
Frontiers in Cardiovascular Medicine 8:806215
2022
A preliminary study of osteochondral regeneration of the femoral medial condyle using a scaffold-free 3D construct of synovial membrane-derived mesenchymal stem cells
Murata D, Ishikawa S, Sunaga T, Saito Y, Sogawa T, Tuji K, Shigenobu K, Nakayama K, Hobo S, Hatazoe T
BMC Veterinary Research 18, 53
2022
Establishment of large canine hepatocyte spheroids by mixing vascular endothelial cells and canine adipose-derived mesenchymal stem cells
Ichikawa A, Neo S, Nukui R, Yamada Y, Nitta S, Iwaki H, Yanagi Y, Nakayama K, Sato S, Tateishi S, Hisasue M
Regenerative Therapy 19, 1–8
2021
Kenzan Method for Scaffold-Free Biofabrication Book
Nakayama K (Ed.)
Springer Nature ISBN 978-3-030-58688-1
2021
Successful use of bio plugs for delayed bronchial closure after pneumonectomy in experimental settings
Moriyama M, Matsumoto K, Taniguchi D, Machino R, Tsuchiya T, Nakayama K, Nagayasu T
Interactive CardioVascular and Thoracic Surgery ivab306
2021
Bio-3D printing iPSCs-derived human chondrocytes for articular cartilage regeneration
Nakamura A, Murata D, Fujimoto R, Tamaki S, Nagata S, Ikeya M, Toguchida J, Nakayama K
Biofabrication 13, 044103
2021
Fabrication of Cardiac Constructs Using Bio-3D Printer Chapter
Arai K, Murata D, Takao S, Nakayama K
In: Pluripotent Stem-Cell Derived Cardiomyocytes, Springer US
2021
Scaffold-based and scaffold-free cardiac constructs for drug testing Review
Arai K, Kitsuka T, Nakayama K
Biofabrication 13, 042001
2021
Long-term Outcome of Sciatic Nerve Regeneration Using Bio3D Conduit Fabricated from Human Fibroblasts in a Rat Sciatic Nerve Model
Ando M, Ikeguchi R, Aoyama T, Tanaka M, Noguchi T, Miyazaki Y, Akieda S, Nakayama K, Matsuda S
Cell Transplantation 30
2021
Bile duct reconstruction using scaffold-free tubular constructs created by Bio-3D Printer
Hamada T, Nakamura A, Soyama A, et al.
Regenerative Therapy 16, 81–89
2021
Study on Pipetting Motion Optimization of Automatic Spheroid Culture System for Spheroid Formation
Shimoto T, Teshima C, Watanabe T, Zhang XY, Ishikawa A, Higaki H, Nakayama K
Journal of Robotics and Mechatronics 33(1), 78–87
2021
Bio-3D Printed Organs as Drug Testing Tools Chapter
Arai K, Nakayama K
In: Kenzan Method for Scaffold-Free Biofabrication, Springer Nature, 149–164
2021
Scaffold-Free Biofabrication for Articular Cartilage (and Subchondral Bone) Chapter
Murata D
In: Kenzan Method for Scaffold-Free Biofabrication, Springer Nature, 149–164
2020
Mechanism of Peripheral Nerve Regeneration Using a Bio 3D Conduit Derived from Normal Human Dermal Fibroblasts
Yurie H, Ikeguchi R, Aoyama T, Ito A, Tanaka M, Oda H, Noguchi T, Takeuchi H, Mitsuzawa S, Ando M, Yoshimoto K, Akieda S, Nakayama K, Matsuda S
J Reconstr Microsurg 37(4), 357–364
2020
Bio 3D Conduits Derived from Bone Marrow Stromal Cells Promote Peripheral Nerve Regeneration
Yurie H, Ikeguchi R, Aoyama T, Tanaka M, Oda H, Takeuchi H, Mitsuzawa S, Ando M, Yoshimoto K, Noguchi T, Akieda S, Nakayama K, Matsuda S
Cell Transplantation 29, 1–9
2020
Pro-angiogenic scaffold-free Bio 3D conduit from human iPSC-derived mesenchymal stem cells promotes peripheral nerve regeneration
Mitsuzawa S, Zhao C, Ikeguchi R, et al.
Scientific Reports 10, 12034
2020
Osteochondral regeneration using adipose tissue derived-mesenchymal stem cells Review
Murata D, Fujimoto R, Nakayama K
Int. J. Mol. Sci. 21(10), 3589
2020
Drug response analysis for scaffold-free cardiac constructs fabricated using bio-3D printer
Arai K, Murata D, Takao S, Nakamura A, Itoh M, Kitsuka T, Nakayama K
Scientific Reports 10, 8972
2020
Osteochondral regeneration using scaffold-free constructs of adipose tissue-derived mesenchymal stem cells made by a bio three-dimensional printer with a needle-array in rabbits
Murata D, Kunitomi Y, Harada K, Tokunaga S, Takao S, Nakayama K
Regenerative Therapy 15, 77–89
2020
Cryopreservation method for spheroids and fabrication of scaffold-free tubular constructs
Arai K, Murata D, Takao S, Verissimo A, Nakayama K
PLoS One 15(4):e0230428
2020
Scaffold-free Bio-3D Printing using Spheroids as “Bio-inks” for Tissue (Re-)construction and Drug Response Tests Review
Murata D, Arai K, Nakayama K
Adv. Healthcare Mater. May 2020
2020
Human lung microvascular endothelial cells as potential alternatives to human umbilical vein endothelial cells in bio-3D-printed trachea-like structures
Taniguchi D, Matsumoto K, Machino R, et al.
Tissue and Cell 63, 101321
2020
A scaffold-free Bio 3D nerve conduit for repair of a 10-mm peripheral nerve defect in the rats
Takeuchi H, Ikeguchi R, Aoyama T, Oda H, Yurie H, Mitsuzawa S, Tanaka M, Ohta S, Akieda S, Miyazaki Y, Nakayama K, Matsuda S
Microsurgery 40(2), 207–216
2019
2-Cl-C.OXT-A Stimulates Contraction through the Suppression of Phosphodiesterase Activity in Human Induced Pluripotent Stem Cell-derived Cardiac Organoids
Kitsuka T, Itoh M, Amamoto S, Arai K, Oyama J, Node K, Toda S, Morita S, Nishida T, Nakayama K
PLoS ONE 14(7):e0213114
2019
The efficacy of a scaffold-free Bio 3D conduit developed from autologous dermal fibroblasts on peripheral nerve regeneration in a canine ulnar nerve injury model
Mitsuzawa S, Ikeguchi R, Aoyama T, Takeuchi H, Yurie H, Oda H, Ohta S, Ushimaru M, Itho T, Tanaka M, Kunitomi Y, Tsuji M, Akieda S, Nakayama K, Matsuda S
Cell Transplantation 28(9-10)
2019
Osteochondral regeneration using constructs of mesenchymal stem cells made by bio three-dimensional printing in mini-pigs
Yamasaki A, Kunitomi Y, Murata D, Sunaga T, Kuramoto T, Sogawa T, Misumi K
J Orthop Res 37(6), 1398–1408
2019
Development of an immunodeficient pig model allowing long-term accommodation of artificial human vascular tubes
Itoh M, Mukae Y, Kitsuka T, et al.
Nature Communications 10, 2244
2019
Histological evaluation of tendon formation using a scaffold-free three-dimensional-bioprinted construct of human dermal fibroblasts under in vitro static tensile culture
Nakanishi Y, Okada T, Takeuchi N, Kozono N, Senjyu T, Nakayama K, Nakashima Y
Regenerative Therapy 11, 47–55
2019
Regenerative medicine using stem cells from human exfoliated deciduous teeth (SHED): a promising new treatment in pediatric surgery
Taguchi T, Yanagi Y, Yoshimaru K, Zhang X, Matsuura T, Nakayama K, Kobayashi K, Yamaza H, Nonaka K, Ohga S
Surgery Today 49(4), 316–322
2019
Regeneration of esophagus using a scaffold-free biomimetic structure created with bio-three-dimensional printing
Takeoka Y, Matsumoto K, Taniguchi D, et al.
PLoS ONE 14(3):e0211339
2019
Replacement of Rat Tracheas by Layered, Trachea-like, Scaffold-free Structures of Human Cells Using Bio-3D Printing
Machino R, Matsumoto K, Taniguchi D, et al.
Adv. Healthcare Mater. 2019, 1800983
2019
A Scaffold-Free Allogeneic Construct From Adipose-Derived Stem Cells Regenerates an Osteochondral Defect in a Rabbit Model
Oshima T, Nakase J, Toratani T, Numata H, Takata Y, Nakayama K, Tsuchiya H
Arthroscopy 35(2), 583–593
2018
Fabrication of scaffold-free tubular cardiac constructs using a bio-3D printer
Arai K, Murata D, Verissimo AR, Mukae Y, Itoh M, Nakamura A, Morita S, Nakayama K
PLoS ONE 13(12):e0209162
2018
The addition of human iPS cell-derived neural progenitors changes the contraction of human iPS cell-derived cardiac spheroids
Mukae Y, Itoh M, Noguchi R, Furukawa K, Arai KI, Oyama JI, Toda S, Nakayama K, Node K, Morita S
Tissue Cell 53, 61–67
2018
Osteochondral regeneration with a scaffold-free three-dimensional construct of adipose tissue-derived mesenchymal stromal cells in pigs
Murata D, Akieda S, Misumi K, Nakayama K
Tissue Engineering and Regenerative Medicine 15(1), 101–113
2018
Scaffold-free trachea regeneration by tissue engineering with bio-three-dimensional printing
Taniguchi D, Matsumoto K, Tsuchiya T, Machino R, Takeoka Y, Elgalad A, Gunge K, Takagi K, Taura Y, Hatachi G, Matsuo N, Yamasaki N, Nakayama K, Nagayasu T
Interactive Cardiovascular and Thoracic Surgery 26(4), 713
2018
Regeneration of diaphragm with bio-3D cellular patch
Zhang X-Y, Yanagi Y, Sheng Z, Nagata K, Nakayama K, Taguchi T
Biomaterials 167, 1–14
2018
A definition of bioinks and their distinction from biomaterial inks Perspective
Groll J, Burdick JA, Cho D-W, ...Nakayama K, et al.
Biofabrication 11, 013001
2017
The efficacy of a scaffold-free bio 3D conduit developed from human fibroblasts on peripheral nerve regeneration in a rat sciatic nerve model
Yurie H, Ikeguchi R, Aoyama T, Kaizawa Y, Tajino J, Ito A, Ohta S, Oda H, Takeuchi H, Akieda S, Tsuji M, Nakayama K, Matsuda S
PLoS One 12(2):e0171448
2017
Scaffold-free tissue-engineered allogenic adipose-derived stem cells promote meniscus healing
Toratani T, Nakase J, Numata H, Oshima T, Takata Y, Tsuchiya H, Nakayama K
Arthroscopy 33(2), 346–354
2017
Principles of the “Kenzan” method for robotic cell spheroid-based three-dimensional bioprinting Review
Moldovan NI, Hibino N, Nakayama K
Tissue Engineering Part B: Reviews 23(3), 237–244
2017
In vivo and ex vivo methods of growing a liver bud through tissue connection
Yanagi Y, Nakayama K, Taguchi T, et al.
Scientific Reports 7, 14085
2017
Scaffold-Free Biofabrication Chapter
Verissimo AR, Nakayama K
In: 3D Printing and Biofabrication, Springer
2016
Development of a three-dimensional pre-vascularized scaffold-free contractile cardiac patch for treating heart disease
Noguchi R, Nakayama K, Itoh M, et al.
J Heart and Lung Transplantation 35(1), 137–145
2015
A preliminary study of osteochondral regeneration using a scaffold-free three-dimensional construct of porcine adipose tissue-derived mesenchymal stem cells
Murata D, Tokunaga S, Tamura T, Kawaguchi H, Miyoshi N, Fujiki M, Nakayama K, Misumi K
J Orthop Surg Res 10, 35
2015
Scaffold-free tubular tissues created by a Bio-3D printer undergo remodeling and endothelialization when implanted in rat aortae
Itoh M, Nakayama K, Noguchi R, et al.
PLoS ONE 10(9):e0136681
2014
Multipotency of equine mesenchymal stem cells derived from synovial fluid
Murata D, Miyakoshi D, Hatazoe T, Miura N, Tokunaga S, Fujiki M, Nakayama K, Misumi K
The Veterinary Journal 202(1), 53–61
2014
Simultaneous regeneration of full-thickness cartilage and subchondral bone defects in vivo using a three-dimensional scaffold-free autologous construct derived from high-density bone marrow-derived mesenchymal stem cells
Ishihara K, Nakayama K, Akieda S, Matsuda S, Iwamoto Y
J Orthop Surg Res 9, 98
2013
In Vitro Biofabrication of Tissues and Organs Chapter
Nakayama K
In: Biofabrication: Micro- and Nano-Fabrication, Printing, Patterning and Assemblies
2013
Bio rapid prototyping project: Development of spheroid formation system for regenerative medicine
Shimoto T, Hidaka N, Sasaki H, Nakayama K, Akieda S, Matsuda S, Miura H, Iwamoto Y
Information Technology Convergence 253, 855–862
2012
Building of HD MACs using cell processing robot for cartilage regeneration
Shimoto T, Nakayama K, Matsuda S, Iwamoto Y
Journal of Robotics and Mechatronics 24(2), 347–353
2009
Mevastatin reduces cartilage degradation in rabbit experimental osteoarthritis through inhibition of synovial inflammation
Akasaki Y, Matsuda S, Nakayama K, Fukagawa S, Miura H, Iwamoto Y
Osteoarthritis Cartilage 17(2), 235–243
2009
Establishment of an animal model of a pasteurized bone graft, with a preliminary analysis of muscle coverage or FGF-2 administration to the graft
Yoshida T, Sakamoto A, Tsukamoto N, Nakayama K, Iwamoto Y
J Orthop Surg Res 4, 31
2005
Contact stress at the post-cam mechanism in posterior-stabilised total knee arthroplasty
Nakayama K, Matsuda S, Miura H, Iwamoto Y, Higaki H, Otsuka K
The Bone & Joint Journal 87-B(4), 483–488
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Center for Regenerative Medicine Research
Faculty of Medicine, Saga University
5-1-1 Nabeshima, Saga City
Saga 849-8501, Japan

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