Biomedical Engineering - Technology for Regenerative Medicine

Cardias Patches

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1 Cardiac patches and bioartificial heart In the United States, heart donor availability has increasingly failed to keep pace with rising demand. According to the United Network for Organ Sharing, 512 patients on the waiting list died in 1988; 527, in 1989; 650, in 19 90, more than 800 in 1991 . T he number and proportion of potential recipients who die awaiting heart transplantation is increasing every year. Congestive heart failure and arrhythmia are the major causes of death . [ 1] Bioartificial heart and cardiac patches seem to be the most promising alternative s to the heart trans plant and to the heart devices (as VAD s , that are devices, only used as a bridge to the transplant). Haneef et al. [2] seeded a 3D collagen scaffold with bone marrow MSCs and then and stimulated them using electrical stimulation to obtain the differentiation into cardiomyocytes, with the final goal to obtain a functional cardiac patch. With this exercise, we want to chara cterize and analyze if this device could be a good candidate as a cardiac patch . A cardiac patch is a cellularised thin scaffold that is a promising therapeutic mode for ventricular wall reconstruction. Bibliography [1] McManus RP , O'Hair DP et al. Patients who die awaiting heart transplantation. J Heart Lung Transplant. 1993 Mar - Apr;12(2):159 - 71. [2] Haneef K., Lila N., et al. Development of bioartificial myocardium by el ectrostimulation of 3D collagen scaffolds seeded with stem cells. Heart International 7, 70 - 76 (2012). [4] Mollova M., Bersell K. et al. Cardiomyocyte proliferation contributes to heart growth in young humans. PNAS Early Edition (2012). [5] Redaelli A., Montevecchi F. Biomeccanica. Analisi multiscala di tessuti biologici, Pàtron (2012). [6] Ott H.C., Matthiesen T.S. et al. Per fusion - decellularized matrix: using nature’s platform to engineer a bioartificial heart. Nat. Med. 14, 213 - 221 ( 2008). [7] Zimmermann W.H. et al. Engineered heart tissue grafts improve systolic and diastolic function in infarcted rat hearts. Nat. Med. 12, 452 – 458 (2006). [8] Radisic M., Deen W. et al. Mathematical model of oxygen distribution in engineered cardiac tissue with parallel channel array perfused with culture medium containing oxygen carriers. Am. J. Physiol. Heart Circ. Physiol.288, H1278 – H1289 (2005). 2 Answer briefly to the que stions in the table What is the therapeutic product? bone marrow MSCs embedded in 3D collagen scaffold What are the elements that identify the therapeutic product as a PTC? The main therapeutic agent are the cells, obtained after "non - minimal manipula tion" consisting in cell isolation and cell expansion and seeding In what phase of the regulatory process for new PTC s can this PTC be localized? in vitro What are the standards that apply in this stage of the PTC process of development? GLP What are the risks associated with this PTC (immunogenic, tumour, teratoma, infection, toxicity)? Immunological rejection : YES the cell sources is unknown Tumor formation YES . It could be due to the collagen scaffold and cell ( low probability from MSCs ) . Teratoma formation: No. Unless they us e P LURIPOTENT STEM cells differentiated in MSC; Transmission of infections YES due to donor mate rial or from any manipulation Administration of toxic contaminants : YES due to donor mate rial or from any manipulation s. 3 Classify all the cell sources potentially usable in human patient s for this therapy Cell source (based on immunogenic ity) Cell type/s Advantages Criticalities Can be used? § (Y/N) Autologous MSC isolated from the bone marrow of the patient and expanded Immunological compatibility. Limited availability. Limited expandability. Invasive test. YES Autologous iPS from somatic cells of the patient, differentiated in MSCs Immunological Compatibility. Limitations in redifferentiation protocols Risk of teratoma NO (terato ma) Syngeneic Embryonic stem cells isolated from clones of the patient and differentiated in MSCs Immunological compatibility with the exception of the mitochondrial DNA. Ethical, technical and regulatory limitations for cloned cells. Limitations in redifferentiation protocols. Risk of teratoma Formation. NO (terato ma) Allogeneic MSCs isolated from a human donor and expanded Available from HLA - matched donors . Can be Industrialized. Immunogenicity. Risk of infection Risk of tumor (it is very limited for MSC) NO (I mmun ogenic it y) Xenogeneic MSCs isolated from a nonhuman donor and expanded Largely available from animal livestock. Can be industrialized. Risk of acute immune rejection. Risk of xeno - zoonoses. Limited biological Functionality. Risk of tumor (limited) NO (acute immun orejecti on and the tumor formati on ) 4 EXERCISES: The scaffolds were composed of lyophilized, non - denatured, native type I collagen (bovine origin) obtained from a commercially available collagen kit (Pangen 2, Urgo Laboratory, Chenove, France). Templates of 25×25×6 mm of collagen scaffol d were placed in 100 mm Petri dishes. A suspension containi ng cardiomyocyte (CM) were seeded on the upper collagen surface with a superficial density of 10 6 cell/cm 2 . The scaffolds were left at 37°C in a 5% CO 2 humidified atmosphere for few hours to allow the cell attachment. Afterwards, 10 mL medium was added and plates were incubated at 37°C in a 5% CO 2 , in order that cells can migrate into the scaffold and proliferate 1) Calculate the time to wait to reach (in the scaffold volume) the cellular density of the human heart, N h = 3.6x10 7 cells/cm 3 . H uman cardiomyocytes doubling time: t d,CM =36 h. SOLUTION 1) Data: Seeding density (It is a superficial density ! ) Ni = 10 6 cell /cm 2 Scaffold volume =25x25x6 mm 3 Final density of cell N h = ! .# ∗ %& ! '())* '+ " t d cm =36h Scheme : S olution: T ime to wait: t = d*t d t d cm =36h Number of division: d= ln(X f /X i)/ln 2 Number of cells per area X=N*A Number of cells per volume X=N*V Number of passages d= ln(X f /X i)/ln 2 Expansion time t = d*t d 5 Initial cell number X i=A s *Ni Seeding Area A s = 25*25=625mm 2 =6.25cm 2 X i=A s *Ni = 6 . 25 '( , %& # '())* '+ $ = 6 . 25 ∗ 10 # ',--. Final number of cells / - = 0 . ∗ 12- * S caffold volume V ol s = 25*25*6mm 3 = 3750mm 3 = 3.75 cm 3 N h = ! .# ∗ %& ! '())* '+ " / - = 0 . ∗ 12- * = 3 . 6 ∗ 10 / ',--. '( ! ∗ 3 . 75 '( ! = 13 . 5 ∗ 10 / ',--. Doublings: 5 = 01 ( %! .3 ∗ %& ! # .,3 ∗ %& # 4 ) 01 , = 4 . 43 passages T otal time 7 67 = 7 8'+ ∗ 5 7 67 = 36ℎ ∗ 4 . 43 = 159 . 48 ℎ ≅ 6 . 6 5