Table 4.
Decellularization studies to generate scaffolds for testicular bioengineering.
| Biomaterial | Protocol of decellulariz ation |
Species involved |
Type of study (in vitro/in vivo) |
Type of Cells Used |
Main biological findings | Reference |
|---|---|---|---|---|---|---|
| Decellularized scaffolds | Via infusion of 0.5% sodium dodecyl sulfate (SDS) for 48 h, followed by 1% Triton X-100 for 6 h and then 1% DNase I for 1 h | Rat | In vitro | Testicular cells | The results revealed that the testis were successfully decellularized while maintaining the three-dimensional structure of the matrix and preserving the extracellular matrix components. After recellularization, the scaffold demonstrated that it supports cell adhesion and proliferation. | (217) |
| Decellularized scaffolds | 0.5% sodium dodecyl sulfate (SDS) and 0.5% (v/v) Triton X-100 were applied for 2 h. | Mouse | In vitro | Spermatogonial Stem cells | After decellularization, the three-dimensional structure and constituents of the ECM remained preserved. The scaffold was successfully recellularized and had good cytocompatibility. In vivo tests showed some specific testicular cells, such as inhibin-positive cells within the scaffolds. In addition, the scaffold provided a microenvironment for DAZL-positive cell migration. | (218) |
| Decellularized scaffolds | Freeze-thaw cycle. After 1% Triton X-100 through the vas deferens for 4 h, 1% SDS for 48 h and 1% DNase for 2 h. | Rat | In vitro e in vivo | Mesenchymal stem cells collected from adult mouse bone marrow | After decellularization, the three-dimensional structure and constituents of the ECM remained preserved. The scaffold was successfully recellularized and had good cytocompatibility, in vivo tests showed some specific testicular cells, such as inhibin-positive cells within the scaffolds. In addition, the scaffold provided a microenvironment for DAZL-positive cell migration. | (60) |
| Decellularized scaffolds | 0.01% sodium dodecyl sulfate for 7 h followed by 1 h of agitation in 1% Triton X-100 | Swine | In vitro | Testicular cell organoids | Testicular cell suspensions isolated from immature porcine testicular tissue can form testicular organoids with seminiferous tubule organization comparable to the native organ when cultured in vitro in hydrogels. Testicular organoids showed somatic cell functionalities that were maintained until the end of the culture. | (43) |
| Decellularized scaffolds | The slices were decellularized in 1% sodium dodecyl sulfate (SDS) and then incubated for 24h. | Ram | In vitro | Spermatogonial Stem cells | The three-dimensional culture of SSCs in decellularized extracellular matrix provided adequate conditions for their preservation and proliferation. The results of this study may be a way to deepen the study of the process of spermatogenesis in vitro, as well as a hope for the treatment of infertility in men. | (219) |
| Decellularized scaffolds | 1% Triton X-100 and/or 1% sodium dodecyl sulfate (SDS) for 24 or 48 hours. | Human | In vitro | Neonatal testicular cells | The scaffolds obtained after decellularization are not cytotoxic, providing adequate conditions that support the fixation and infiltration of testicular cells. | (220) |
| Decellularized scaffolds | Concentrations of 1%, 0.1% and 0.01% SDS were tested in the SDS-Triton (ST) and Triton-SDS-Triton (TST) protocols. | Swine | In vitro | Human primary Sertoli cells | The conditions of 0.1% and 3% TET offered the best decellularization in terms of DNA elimination and extracellular matrix (ECM) preservation, ensuring good fixation, proliferation and functionality of human Sertoli cells. | (221) |
| Decellularized scaffolds | 0.5% (v/v) sodium dodecyl sulfate diluted in distilled water for 18 hours + 0.5% (v/v) Triton X-100 diluted in distilled water for 18 hours. | Mouse | In vitro | Spermatogonial Stem cells | Treatment of the mice’s whole testis with Triton X-100 and SDS efficiently removed the cells from the testis, so it is an appropriate protocol for the decellularization of whole testis. | (222) |
| Decellularized scaffolds | SDS | Rat | In vitro | Embryoid bodies | Recellularized testicular ECM may be a promissing tool for future new approaches for testicular cell differentiation applied for assisted reproduction techniques and infertility treatments | (223) |
| Decellularized scaffolds | 100μm slices were decellularized with 1% SDS immersed for 24 hours. | Ram | In vitro | Spermatogonial stem cells | The results of the present study indicated that testicular scaffolds provide adequate conditions for the differentiation of SSCs. | (224) |
| Decellularized scaffolds | 1% sodium dodecyl sulfate (SDS) in PBS solution for 18 h. | Mouse | In vitro | Mouse spermatogonial stem cells | The hydrogel scaffold containing 10 mg/ml decellularized ECM maintained the properties of SSCs at the molecular and cellular levels and promoted the differentiation of SSCs into round spermatids in the absence of somatic cells. | (58) |
| Decellularized scaffolds | Sodium dodecyl sulfate (SDS) 0.5%, 1%, 2%, Trypsin-EDTA 0.5%, 1%, Triton X-100 1% and 2%, respectively. | Ram | – | – | The 1% SDS perfusion protocol for 6-8 hours generated an acellular scaffold maintaining the integrity of the vascular network and preserving the three-dimensional structure as well as the extracellular matrix components. | (225) |
| Decellularized scaffolds | Hypertonic tris-buffer (TBS), 50 mM Tris-HCl pH 7.6 for 30 min, followed by 0.1% Triton X-100 for 15 min. | Ram | In vitro | Mouse spermatogonial stem cells | When cultured in acellular scaffolds, neonatal testicular cells from mice produced morphologically mature sperm in the shortest possible time. The scaffolds provided a microenvironment that functionally supported testicular cells, which secreted testosterone and inhibin B. | (179) |
| Decellularized scaffolds | 1: 0.1% SDS for 24 hours 2: 0.5% SDS for 24 hours. 3: 1% SDS for 24 hours. 4: 0.5% SDS for 18 hours, then washed with PBS and immersed in 0.5% Triton for 18 hours. |
Rat | In vitro | Spermatogonial cells | Immersion of testis from adult mice in 0.5% SDS solution and 0.5% triton solution was an effective method for decellularization of whole testicles without damaging the seminiferous tubules. The decellularized testicular scaffolds were biocompatible and had no detrimental effect on the viability of spermatogonial cells. Generated scaffolds supported spermatogonial cell proliferation during two weeks of culture. | (226) |
| Decellularized scaffolds | 0.5% (v/v) Sodium dodecyl sulfate, then in 0.5% (v/v) Triton X-100, for 18 hours. | Rat | In vitro | Spermatogonial stem cells (6-day-old) | Spermatogonial stem cells can proliferate and differentiate into spermatocytes after being injected into decellularized testicular structures. | (62) |
| Decellularized scaffolds | Sodium hypochlorite solution 1.25% | Human | In vitro e in vivo | Induced Pluripotent Stem cells (iPS) | A 3D cell culture model was developed to generate human male germ cells from iPSCs and this model was compared to conventional 2D culture. Considering the effect of the 3D scaffold in the induction of specific markers of male germ cells, an increase in the efficiency of germ cell differentiation was observed. | (1) |