We investigated whether human testis-derived cells (hTCs) obtained by testicular sperm extraction (TESE) can be cultured on a homemade scaffold composed of electrospun nanofibers of homogeneous poly (vinyl alcohol)/human serum albumin/gelatin (PVA/HSA/gelatin)
September 8, 2021
We investigated whether human testis-derived cells (hTCs) obtained by testicular sperm extraction (TESE) can be cultured on a homemade scaffold composed of electrospun nanofibers of homogeneous poly (vinyl alcohol)/human serum albumin/gelatin (PVA/HSA/gelatin). Materials and Methods In this experimental lab study, human TCs underwent two actions of enzymatic cell isolation and five Rilmenidine Phosphate culture passages. hematoxylin and eosin (H&E) staining and SEM. Cell viability study using MTT [3-(4, 5-dimethyl-2-thiazolyl) -2, 5-diphenyl -2H- tetrazolium bromide] assay was performed on days 7 and 14. Results Visualization by H&E staining and SEM indicated that hTCs were seeded around the scaffold. MTT test showed that this PVA/HSA/gelatin scaffold is not toxic for hTCs. Conclusion It seems that this PVA/HSA/gelatin scaffold is usually supportive for growth of hTCs. Keywords: Azoospermia, Human Serum Albumin, Scaffold, Testis, Tissue Engineering Introduction Almost DPP4 7% of all men, including those who lack spermproduction, suffer from infertility (1). Stem cells, with theirgreat and unique capacity to form other cell types, have raisedhuge hopes for scientists and clinicians as well as patientswith male infertility. After the derivation of mouse embryonicstem cells (2), different studies used mouse primordial germcells (PGCs) to investigate the biology of germ cells and theirprogenitors (3, 4). Later, in 1998, the first pluripotent stemcells were generated from pre-implantation human embryosin blastocyst stage (5), and also human PGCs (6) which werenamed human embryonic stem cells (hESCs) and humanembryonic germ cells (hEGCs), respectively. Since 2003, several studies have shown the potential of the ESCs to formmale and female germ cells (7-10). However, no gametehas been produced so far. Some investigations made effortsto reprogram unipotent spermatogonial stem cells (SSCs) to derive pluripotent germ-line stem cells (GSCs) in vitro in mice (11), rats (12) and humans (13). Nonetheless, laterreports indicated that human testis-derived cells (hTCs) are not pluripotent and possess characteristics similar to those of mesenchymal stromal cells (14). The latest studies includingIrie and Suranis investigations (15) revealed that germ celldevelopment in humans differs from that in mice especiallyin terms of gene expression profile, which might be thereason for variations in results. Despite improvements inthe field, there are still challenges for translation of stem cell biotechnology to bedside practice (i.e. has not yet been used in male reproductive/regenerative medicine). In a recent study, mouse fertile sperm production from GSCs was done using organ culture (16). Besides developmental differences between mouse and human germ cells, there are more restrictive ethical issues regarding human organ culture compared to mouse organ culture. Therefore, tissue engineering methods are highly required for regeneration of some tissues and organs. These methods prepare bio-scaffolds to promote the development of new tissues such as cartilage or bone. In comparison with other instances where tissue engineering produced artificial tissues, researchers in the field of human male infertility Rilmenidine Phosphate could not obtain adequate mature cells. In regenerative medicine, utilization of pluripotent or multipotent stem cells has higher chance of success compared to unipotent cells like human SSCs (17). We previously showed the mulipotency of hTCs obtained from TESE samples (14). The aim of this study was to make a homemade scaffold composed of electrospun fibers of homogeneous solution of poly (vinyl alcohol)/ human serum albumin/gelatin (PVA/HSA/gelatin) as a niche for hTCs. Development of an artificial organ culture for production of male germ cells from hESC-derived GSCs could be the ultimate goal in this field. Materials and Methods Fabrication of the scaffold In this experimental lab study, initially 450 mg of PVA powder (Merck, Germany, MW 72,000) was dissolved in deionized water (to reach a final concentration of 7% w/v) in a final volume of 6 mL which was kept at 80C for 5 hours in a sterile beaker to Rilmenidine Phosphate make a clear solution. Next, 0.3 g gelatin powder (Merck, Germany) was added and the mixture was mixed by a magnetic stirrer at room temperature (RT). Then, 2 mL of a 20 g/dL solution of HSA (CSL Behring AG, Switzerland) was added to the mixture and mixed for 60 minutes on a magnetic stirrer. The resulting solution was.