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1
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Ghorbani M, Hassani R, Nourani MR, Goodarzi V. Advanced 3Dimentional engineered microenvironment to improve of in vitro spermatogenesis: narrative review. JBRA Assist Reprod 2025; 29:160-166. [PMID: 39835797 PMCID: PMC11867261 DOI: 10.5935/1518-0557.20240077] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2024] [Accepted: 12/30/2024] [Indexed: 01/22/2025] Open
Abstract
Induction of in vitro spermatogenesis may be helpful in the treatment of infertility in azoospermic individuals and those undergoing chemotherapy. Different cultivation systems have been implemented to achieve this aim. This review study aimed to investigate the application of three-dimensional culture in the induction of in vitro spermatogenesis. Relevant studies published in English were identified using PubMed using a range of search terms related to the core focus on tissue engineering of male reproductive systems, in vitro spermatogenesis, germ cell preservation, 3D culture systems for in vitro spermatogenesis, a 3D culture of testis tissue with were last updated in end of 2023. Searches were not restricted to a particular time frame or species, although the emphasis within the review is on regenerative medicine in mammalian male fertility preservation and in vitro spermatogenesis. Spermatogenesis is one of the most complicated cellular differentiation processes in the body. Significant attempts have been made to control spermatogenesis to drive differentiation of male germ stem cells toward mature sperm. Current research efforts focus on providing appropriate microenvironmental conditions to support the process of in vitro spermatogenesis by applying the principles of cell transplantation, material science, and bioengineering. Regenerative medicine may open a new avenue to patients for restoration and maintenance of normal function in spermatogenesis. The techniques reviewed are still in development, and this paper can become the primary reference for a large body of scientists developing advanced tissue engineering for male germ cells or developing the next generation of reproductive medicine.
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Affiliation(s)
- Masoud Ghorbani
- Tissue Engineering and Regenerative Medicine Research Center,
Baqiyatallah University of Medical Sciences, Tehran, Iran
| | - Roya Hassani
- Department of Anatomy, School of Medicine, Iran University of
Medical Sciences, Tehran, Iran
| | - Mohammad Reza Nourani
- Tissue Engineering and Regenerative Medicine Research Center,
Baqiyatallah University of Medical Sciences, Tehran, Iran
| | - Vahabodin Goodarzi
- Tissue Engineering and Regenerative Medicine Research Center,
Baqiyatallah University of Medical Sciences, Tehran, Iran
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2
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Huang R, Xia H, Meng T, Fan Y, Tang X, Li Y, Zhang T, Deng J, Yao B, Huang Y, Yang Y. Construction of human pluripotent stem cell-derived testicular organoids and their use as humanized testis models for evaluating the effects of semaglutide. Theranostics 2025; 15:2597-2623. [PMID: 39990223 PMCID: PMC11840739 DOI: 10.7150/thno.104523] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2024] [Accepted: 01/15/2025] [Indexed: 02/25/2025] Open
Abstract
Background: The generation of human testicular organoids from human induced pluripotent stem cells (hiPSCs) presents exciting opportunities for gonadal developmental biology, and reproductive disease modeling. However, creating organoids that closely mimic the tissue structure of testes remains challenging. Methods: In this study, we established a method for generating testicular organoids (TOs) from hiPSCs using a stepwise differentiation approach and a combination of hanging drop and rotational culture systems. The capability of hiPSC-derived precursor testicular cells to self-assemble into organoids was confirmed by detection of morphology, single-cell RNA-sequencing, and protein profiles. The reliability of testicular organoids as a drug evaluation model was assessed by the measurements of transcriptome signatures and functional features, including hormone responsiveness and blood-testis barrier (BTB) formation, and drug sensitivity assessment by recording cell viability and BTB integrity in organoids exposed to reproductive toxicants. Finally, we applied testicular organoids to evaluate the effects of semaglutide, a glucagon-like peptide-1 receptor agonist (GLP-1 RA), on testicular function, thereby underscoring their utility as a model for drug evaluation. Results: These organoids exhibited testicular cord-like structures and BTB function. RNA sequencing and functional assays confirmed that testicular organoids possess gene expression profiles and endocrine functions regulated by gonadotropins, closely resembling those of testicular tissue. Notably, these organoids displayed sensitivity to semaglutide. Treatment with semaglutide resulted in reduced testosterone levels and downregulation of INHBB expression, aligning with previous clinical observations. Conclusions: These findings introduced a method for generating testicular organoids from human pluripotent stem cells, highlighting their potential as valuable models for studying testicular function, drug toxicity, and the effects of compounds like semaglutide on testicular health.
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Affiliation(s)
- Rufei Huang
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Huan Xia
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Tao Meng
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Yufei Fan
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Xun Tang
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Yifang Li
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Tiantian Zhang
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Jingxian Deng
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Bing Yao
- Department of Reproductive Medicine, Jinling Hospital, Affiliated Hospital of Medical School, Nanjing University, Nanjing, 210002, China
| | - Yadong Huang
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
- National Engineering Research Center of Genetic Medicine, Guangzhou, 510632, China
- Guangdong Province Key Laboratory of Bioengineering Medicine, Guangzhou, 510632, China
| | - Yan Yang
- State Key Laboratory of Bioactive Molecules and Druggability Assessment, Guangdong Basic Research Center of Excellence for Natural Bioactive Molecules and Discovery of Innovative Drugs, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
- National Engineering Research Center of Genetic Medicine, Guangzhou, 510632, China
- Guangdong Province Key Laboratory of Bioengineering Medicine, Guangzhou, 510632, China
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Tan J, Li J, Lin C, Ye N, Zhang H, Liu C, Han S, Li Z, Zhou X. Generation of mouse testicular organoids with highly compartmentalized tubular lumen structure and their cryopreservation. Life Sci 2024; 355:122980. [PMID: 39147312 DOI: 10.1016/j.lfs.2024.122980] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/24/2024] [Revised: 07/29/2024] [Accepted: 08/12/2024] [Indexed: 08/17/2024]
Abstract
Testicular organoids have great potential for maintaining male fertility and even restoring male infertility. However, existing studies on generating organoids with testis-specific structure and function are scarce and come with many limitations. Research on cryopreservation of testicular organoids is even more limited, and inappropriate cryopreservation methods may result in the loss of properties in resuscitated or regenerated organoids, rendering them unsuitable for clinical or research needs. In this paper, we investigated the effects of mouse age and cell number on the self-aggregation of testicular cells into spheres in low-adsorption plates. Various media compositions, culture systems, and cell numbers were used to culture cell spheres for 14 days to form testicular organoids, and the self-organization of the organoids was assessed by histological and immunofluorescence staining. We determined the appropriate cryopreservation conditions for testicular cells, cell spheres, and tissues. Subsequently, organoids derived from cryopreserved testicular tissues, testicular cells, and testicular cell spheres were compared and evaluated by histological and immunofluorescence staining. The results indicate that testicular cell spheres consisting of 30 × 104 testicular cells from 2-week-old mice were able to form organoids highly similar to the luminal structure and cell distribution of natural mouse testicular tissues. This transformation occurred over 14 days of incubation in α-MEM medium containing 10 % knockout serum replacer (KSR) using an agarose hydrogel culture system. Additionally, the Sertoli cells were tightly connected to form a blood-testis barrier. The relative rates of tubular area, germ cells, Sertoli cells, and peritubular myoid cells were 36.985 % ± 0.695, 13.347 % ± 3.102, 47.570 % ± 0.379, and 27.406 % ± 1.832, respectively. The optimal cryopreservation protocol for primary testicular cells involved slow freezing with a cryoprotectant consisting of α-MEM with 10 % dimethyl sulfoxide (DMSO). Slow freezing with cryoprotectants containing 5 % DMSO and 5 % ethylene glycol (EG) was optimal for all different volumes of testicular cell spheres. Compared to testicular organoids generated from frozen testicular tissue and cell spheres, freezing testicular cells proved most effective in maintaining organoid differentiation characteristics and cell-cell interactions. The findings of this study contribute to a "universal" testicular organoid in vitro culture protocol with promising applications for fertility preservation and restoration in prepubertal cancer patients and adult infertile patients.
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Affiliation(s)
- Jia Tan
- Institute of Biomedical Technology, University of Shanghai for Science and Technology, Shanghai 200093, China
| | - Jiahui Li
- Institute of Biomedical Technology, University of Shanghai for Science and Technology, Shanghai 200093, China
| | - Chunyan Lin
- Institute of Biomedical Technology, University of Shanghai for Science and Technology, Shanghai 200093, China
| | - Na Ye
- Institute of Biomedical Technology, University of Shanghai for Science and Technology, Shanghai 200093, China
| | - Hui Zhang
- Institute of Biomedical Technology, University of Shanghai for Science and Technology, Shanghai 200093, China
| | - Chenxi Liu
- Institute of Biomedical Technology, University of Shanghai for Science and Technology, Shanghai 200093, China
| | - Sha Han
- Department of Andrology, The Center for Men's Health, Urologic Medical Center, Shanghai Key Laboratory of Reproductive Medicine, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200080, China
| | - Zheng Li
- Department of Andrology, The Center for Men's Health, Urologic Medical Center, Shanghai Key Laboratory of Reproductive Medicine, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200080, China
| | - Xinli Zhou
- Institute of Biomedical Technology, University of Shanghai for Science and Technology, Shanghai 200093, China.
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4
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Cham TC, Ibtisham F, Al-Dissi A, Honaramooz A. An in vitro testicular organoid model for the study of testis morphogenesis, somatic cell maturation, endocrine function, and toxicological assessment of endocrine disruptors. Reprod Toxicol 2024; 128:108645. [PMID: 38897308 DOI: 10.1016/j.reprotox.2024.108645] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2024] [Revised: 06/12/2024] [Accepted: 06/13/2024] [Indexed: 06/21/2024]
Abstract
Male reproductive capacity has fallen considerably in recent decades; in addition, the incidence of testicular cancer has increased in many developed countries. The cause of this phenomenon is unknown, but environmental toxicants are considered a major contributing factor. To study potential reproductive toxicants, robust in vitro testis models are needed. We have recently established a porcine testis organoid system with a high resemblance to the architectures of innate testis tissue. Here, we further investigated the testis morphogenesis, cell maturation, and endocrine function of the testis organoids. We also challenged this system with abiraterone, a steroidogenic inhibitor, to validate its suitability as an in vitro platform for endocrine toxicology tests. Our results showed that the testis cells in the organoids reorganize into testis cordal structures, and the cordal relative areas increase in the organoids over time of culture. Moreover, the diameters and cell numbers per cross-section of the cordal structures increased over time. Interestingly, Sertoli cells in the organoids gradually underwent maturational changes by showing increased expression of androgen receptors, decreased expression of the anti-müllerian hormone, and formation of the blood-testis barrier. Next, we confirmed that the organoids respond to hormonal stimulation and release multiple sex hormones, including testosterone, estradiol, and progesterone. Finally, we showed that the production of testosterone and estradiol in this system can be inhibited in response to the steroidogenic inhibitor. Taken together, our organoid system provides a promising in vitro platform for male reproductive toxicology studies on testis morphogenesis, somatic cell maturation, and endocrine production.
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Affiliation(s)
- Tat-Chuan Cham
- Department of Veterinary Biomedical Sciences, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, SK S7N 5B4, Canada
| | - Fahar Ibtisham
- Department of Veterinary Biomedical Sciences, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, SK S7N 5B4, Canada
| | - Ahmad Al-Dissi
- Department of Veterinary Pathology, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, SK S7N 5B4, Canada
| | - Ali Honaramooz
- Department of Veterinary Biomedical Sciences, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, SK S7N 5B4, Canada.
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5
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Zhang Y, Liu K, He H, Xiao H, Fang Z, Chen X, Li H. Innovative explorations: unveiling the potential of organoids for investigating environmental pollutant exposure. ENVIRONMENTAL SCIENCE AND POLLUTION RESEARCH INTERNATIONAL 2024; 31:16256-16273. [PMID: 38342830 DOI: 10.1007/s11356-024-32256-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/21/2023] [Accepted: 01/25/2024] [Indexed: 02/13/2024]
Abstract
As the economy rapidly develops, chemicals are widely produced and used. This has exacerbated the problems associated with environmental pollution, raising the need for efficient toxicological evaluation techniques to investigate the toxic effects and mechanisms of toxicity of environmental pollutants. The progress in the techniques of cell culture in three dimensions has resulted in the creation of models that are more relevant in terms of biology and physiology. This enables researchers to study organ development, toxicology, and drug screening. Adult stem cells (ASCs) and induced pluripotent stem cells (iPSCs) can be obtained from various mammalian tissues, including cancerous and healthy tissues. Such stem cells exhibit a significant level of tissue memory and ability to self-assemble. When cultivated in 3D in vitro environments, the resulting organoids demonstrate a remarkable capacity to recapitulate the cellular composition and function of organs in vivo. Recently, many tumors' tissue-derived organoids have been widely used in research on tumor pathogenesis, drug development, precision medicine, and other fields, including those derived from colon cancer, cholangiocarcinoma, liver cancer, and gastric cancer. However, the application of organoid models for evaluating the toxicity of environmental pollutants is still in its infancy. This review introduces the characteristics of the toxicity responses of organoid models upon exposure to pollutants from the perspectives of organoid characteristics, tissue types, and their applications in toxicology; discusses the feasibility of using organoid models in evaluating the toxicity of pollutants; and provides a reference for future toxicological studies on environmental pollutants based on organoid models.
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Affiliation(s)
- Yuanhang Zhang
- School of Environment, Nanjing Normal University, Nanjing, 210023, China
| | - Kai Liu
- School of Environment, Nanjing Normal University, Nanjing, 210023, China
| | - Huan He
- School of Environment, Nanjing Normal University, Nanjing, 210023, China
- Jiangsu Province Engineering Research Center of Environmental Risk Prevention and Emergency Response Technology, Nanjing, 210023, China
| | - Hui Xiao
- School of Environment, Nanjing Normal University, Nanjing, 210023, China
| | - Zhihong Fang
- School of Environment, Nanjing Normal University, Nanjing, 210023, China
| | - Xianxian Chen
- School of Environment, Nanjing Normal University, Nanjing, 210023, China
| | - Huiming Li
- School of Environment, Nanjing Normal University, Nanjing, 210023, China.
- Jiangsu Province Engineering Research Center of Environmental Risk Prevention and Emergency Response Technology, Nanjing, 210023, China.
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6
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Kimmins S, Anderson RA, Barratt CLR, Behre HM, Catford SR, De Jonge CJ, Delbes G, Eisenberg ML, Garrido N, Houston BJ, Jørgensen N, Krausz C, Lismer A, McLachlan RI, Minhas S, Moss T, Pacey A, Priskorn L, Schlatt S, Trasler J, Trasande L, Tüttelmann F, Vazquez-Levin MH, Veltman JA, Zhang F, O'Bryan MK. Frequency, morbidity and equity - the case for increased research on male fertility. Nat Rev Urol 2024; 21:102-124. [PMID: 37828407 DOI: 10.1038/s41585-023-00820-4] [Citation(s) in RCA: 19] [Impact Index Per Article: 19.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 08/31/2023] [Indexed: 10/14/2023]
Abstract
Currently, most men with infertility cannot be given an aetiology, which reflects a lack of knowledge around gamete production and how it is affected by genetics and the environment. A failure to recognize the burden of male infertility and its potential as a biomarker for systemic illness exists. The absence of such knowledge results in patients generally being treated as a uniform group, for whom the strategy is to bypass the causality using medically assisted reproduction (MAR) techniques. In doing so, opportunities to prevent co-morbidity are missed and the burden of MAR is shifted to the woman. To advance understanding of men's reproductive health, longitudinal and multi-national centres for data and sample collection are essential. Such programmes must enable an integrated view of the consequences of genetics, epigenetics and environmental factors on fertility and offspring health. Definition and possible amelioration of the consequences of MAR for conceived children are needed. Inherent in this statement is the necessity to promote fertility restoration and/or use the least invasive MAR strategy available. To achieve this aim, protocols must be rigorously tested and the move towards personalized medicine encouraged. Equally, education of the public, governments and clinicians on the frequency and consequences of infertility is needed. Health options, including male contraceptives, must be expanded, and the opportunities encompassed in such investment understood. The pressing questions related to male reproductive health, spanning the spectrum of andrology are identified in the Expert Recommendation.
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Affiliation(s)
- Sarah Kimmins
- Department of Pharmacology and Therapeutics, Faculty of Medicine, McGill University, Montreal, Quebec, Canada
- The Centre de Recherche du Centre Hospitalier de l'Université de Montréal, Montreal, Quebec, Canada
- The Département de Pathologie et Biologie Cellulaire, Université de Montréal, Montreal, Quebec, Canada
| | - Richard A Anderson
- MRC Centre for Reproductive Health, University of Edinburgh, Edinburgh, UK
| | - Christopher L R Barratt
- Division of Systems Medicine, School of Medicine, Ninewells Hospital and Medical School, University of Dundee, Dundee, UK
| | - Hermann M Behre
- Center for Reproductive Medicine and Andrology, University Hospital, Martin Luther University Halle-Wittenberg, Halle, Germany
| | - Sarah R Catford
- Hudson Institute of Medical Research, Melbourne, Victoria, Australia
- Department of Obstetrics and Gynaecology, The Royal Women's Hospital, Melbourne, Victoria, Australia
| | | | - Geraldine Delbes
- Institut National de la Recherche Scientifique, Centre Armand-Frappier Sante Biotechnologie, Laval, Quebec, Canada
| | - Michael L Eisenberg
- Department of Urology and Obstetrics and Gynecology, Stanford University, Stanford, CA, USA
| | - Nicolas Garrido
- IVI Foundation, Instituto de Investigación Sanitaria La Fe, Valencia, Spain
| | - Brendan J Houston
- School of BioSciences and Bio21 Institute, The University of Melbourne, Parkville, Melbourne, Australia
| | - Niels Jørgensen
- Department of Growth and Reproduction, International Center for Research and Research Training in Endocrine Disruption of Male Reproduction and Child Health, Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark
| | - Csilla Krausz
- Department of Experimental and Clinical Biomedical Sciences, 'Mario Serio', University of Florence, University Hospital of Careggi Florence, Florence, Italy
| | - Ariane Lismer
- Department of Pharmacology and Therapeutics, Faculty of Medicine, McGill University, Montreal, Quebec, Canada
| | - Robert I McLachlan
- Hudson Institute of Medical Research and the Department of Obstetrics and Gynaecology, Monash University, Melbourne, Australia
- Monash IVF Group, Richmond, Victoria, Australia
| | - Suks Minhas
- Department of Surgery and Cancer Imperial, London, UK
| | - Tim Moss
- Healthy Male and the Department of Obstetrics and Gynaecology, Monash University, Melbourne, Victoria, Australia
| | - Allan Pacey
- Faculty of Biology, Medicine and Health, University of Manchester, Manchester, UK
| | - Lærke Priskorn
- Department of Growth and Reproduction, International Center for Research and Research Training in Endocrine Disruption of Male Reproduction and Child Health, Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark
| | - Stefan Schlatt
- Centre for Reproductive Medicine and Andrology, University of Münster, Münster, Germany
| | - Jacquetta Trasler
- Departments of Paediatrics, Human Genetics and Pharmacology & Therapeutics, McGill University and Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada
| | - Leonardo Trasande
- Center for the Investigation of Environmental Hazards, Department of Paediatrics, NYU Grossman School of Medicine, New York, NY, USA
| | - Frank Tüttelmann
- Institute of Reproductive Genetics, University of Münster, Münster, Germany
| | - Mónica Hebe Vazquez-Levin
- Instituto de Biología y Medicina Experimental, Consejo Nacional de Investigaciones Científicas y Técnicas de Argentina, Fundación IBYME, Buenos Aires, Argentina
| | - Joris A Veltman
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK
| | - Feng Zhang
- Obstetrics and Gynecology Hospital, Institute of Reproduction and Development, Fudan University, Shanghai, China
| | - Moira K O'Bryan
- School of BioSciences and Bio21 Institute, The University of Melbourne, Parkville, Melbourne, Australia.
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7
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Nikmahzar A, Khadivi F, Koruji M, Jahanshahi M, Dehghan Tarazjani M, Shabani M, Abbasi Y, Abbasi M. Evaluation of Apoptosis-related Genes and Hormone Secretion Profiles Using Three Dimensional Culture System of Human Testicular Organoids. Galen Med J 2023; 12:e2805. [PMID: 38774852 PMCID: PMC11108674 DOI: 10.31661/gmj.v12i0.2805] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2022] [Revised: 12/25/2022] [Accepted: 01/21/2023] [Indexed: 05/24/2024] Open
Abstract
BACKGROUND In reproductive biology, testicular organoids can be used to treat infertility and to study testicular development and spermatogonial stem cells (SSCs) differentiation. Generating organoid from primary cells is challenging. In this study, testicular organoids were created using human primary testicular cells and evaluated the apoptotic gene expression and hormone secretion profiles of the organoids. MATERIALS AND METHODS Primary human testicular cells were isolated using 2-step enzymatic digestion from three brain-dead donors. Immunocytochemistry and flow cytometry analyses were performed to confirm human SSCs. Isolated cells were cultured in three experimental groups: control group (2 dimensional (2D)), group 1 (organoid culture after 2D culture), and group 2 (organoid culture immediately after enzymatic digestion). Testicular organoids were cultured in DMEM/F-12 media supplemented with follicle-stimulating hormone (FSH) and fetal bovine serum (FBS) for four weeks. After 24 hours and four weeks of culture, reverse transcription quantitative real-time PCR (RT-qPCR) was used to investigate the relative expression of apoptotic genes (caspase 3, 9, Bax, and Bcl-2). At 24 hours, two weeks, and four weeks after culture, enzyme-linked immunoassay (ELISA) was used to determine the testosterone and inhibin B concentrations. Light microscopy and toluidine blue staining were also used for morphological analysis. RESULTS RT-qPCR results revealed that pro-apoptotic (caspase 3, 9, Bax) gene expression levels were highest in group 2 after 24 h and four weeks of culture. In contrast, the expression level of Bcl-2 (anti-apoptotic) was lower in group 2 compared to other groups. The hormone secretion levels decreased in a time-dependent manner during the cultivation. According to morphological evaluations, testicular organoids are compact, spherical structures with two to three elongated cells organized along their border. CONCLUSION Our findings revealed that the testicular organoid culture system maintained hormonal secretory abilities, demonstrating the function of Sertoli and Leydig cells in the absence of testis-specific environments.
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Affiliation(s)
- Aghbibi Nikmahzar
- Department of Anatomy, School of Medicine, Tehran University of Medical Sciences,
Tehran, Iran
| | - Farnaz Khadivi
- Department of Anatomy, School of Medicine, Shahrekord University of Medical
Sciences, Shahrekord, Iran
| | - Morteza Koruji
- Stem Cell and Regenerative Center, Iran University of Medical Sciences, Tehran, Iran
- Department of Anatomy, School of Medicine, Iran University of Medical Sciences,
Tehran, Iran
| | - Mehrdad Jahanshahi
- Neuroscience Research Center, Department of Anatomy, Faculty of Medicine, Golestan
University of Medical Sciences, Gorgan, Iran
| | - Masoomeh Dehghan Tarazjani
- Vali-E-Asr Reproductive Research Center, Family Research Institute, Tehran
University of Medical Sciences, Tehran, Iran
| | - Maryam Shabani
- Department of Anatomy, School of Medicine, Tehran University of Medical Sciences,
Tehran, Iran
| | - Yasaman Abbasi
- Program in Neuroscience, Center to Advance Chronic Pain Research, Department of
Neural and Pain Sciences, School of Dentistry, University of Maryland, Baltimore,
MD, United States
| | - Mehdi Abbasi
- Department of Anatomy, School of Medicine, Tehran University of Medical Sciences,
Tehran, Iran
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8
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Horvath-Pereira BDO, Almeida GHDR, da Silva Júnior LN, do Nascimento PG, Horvath Pereira BDO, Fireman JVBT, Pereira MLDRF, Carreira ACO, Miglino MA. Biomaterials for Testicular Bioengineering: How far have we come and where do we have to go? Front Endocrinol (Lausanne) 2023; 14:1085872. [PMID: 37008920 PMCID: PMC10060902 DOI: 10.3389/fendo.2023.1085872] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/31/2022] [Accepted: 02/24/2023] [Indexed: 03/18/2023] Open
Abstract
Traditional therapeutic interventions aim to restore male fertile potential or preserve sperm viability in severe cases, such as semen cryopreservation, testicular tissue, germ cell transplantation and testicular graft. However, these techniques demonstrate several methodological, clinical, and biological limitations, that impact in their results. In this scenario, reproductive medicine has sought biotechnological alternatives applied for infertility treatment, or to improve gamete preservation and thus increase reproductive rates in vitro and in vivo. One of the main approaches employed is the biomimetic testicular tissue reconstruction, which uses tissue-engineering principles and methodologies. This strategy pursues to mimic the testicular microenvironment, simulating physiological conditions. Such approach allows male gametes maintenance in culture or produce viable grafts that can be transplanted and restore reproductive functions. In this context, the application of several biomaterials have been proposed to be used in artificial biological systems. From synthetic polymers to decellularized matrixes, each biomaterial has advantages and disadvantages regarding its application in cell culture and tissue reconstruction. Therefore, the present review aims to list the progress that has been made and the continued challenges facing testicular regenerative medicine and the preservation of male reproductive capacity, based on the development of tissue bioengineering approaches for testicular tissue microenvironment reconstruction.
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Affiliation(s)
| | | | | | - Pedro Gabriel do Nascimento
- Department of Surgery, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo, Brazil
| | | | | | | | - Ana Claudia Oliveira Carreira
- Department of Surgery, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo, Brazil
- Centre for Natural and Human Sciences, Federal University of ABC, São Paulo, Brazil
| | - Maria Angelica Miglino
- Department of Surgery, School of Veterinary Medicine and Animal Science, University of São Paulo, São Paulo, Brazil
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9
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Patrício D, Santiago J, Mano JF, Fardilha M. Organoids of the male reproductive system: Challenges, opportunities, and their potential use in fertility research. WIREs Mech Dis 2023; 15:e1590. [PMID: 36442887 DOI: 10.1002/wsbm.1590] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/02/2022] [Revised: 10/17/2022] [Accepted: 11/12/2022] [Indexed: 11/30/2022]
Abstract
Organoids are units of function of a given organ able to reproduce, in culture, a biological structure similar in architecture and function to its counterpart in vivo. Today, it is possible to develop an organoid from a fragment of tissue, a stem cell located in an adult organ, an embryonic stem cell, or an induced pluripotent stem cell. In the past decade, many organoids have been developed which mimic stomach, pancreas, liver and brain tissues, optic cups, among many others. Additionally, different male reproductive system organs have already been developed as organoids, including the prostate and testis. These 3D cultures may be of great importance for urological cancer research and have the potential to be used in fertility research for the study of spermatozoa production and maturation, germ cells-somatic cells interactions, and mechanisms of disease. They also provide an accurate preclinical pipeline for drug testing and discovery, as well as for the study of drug resistance. In this work, we revise the current knowledge on organoid technology and its use in healthcare and research, describe the male reproductive system organoids and other biomaterials already developed, and discuss their current application. Finally, we highlight the research gaps, challenges, and opportunities in the field and propose strategies to improve the use of organoids for the study of male infertility situations. This article is categorized under: Reproductive System Diseases > Stem Cells and Development Reproductive System Diseases > Biomedical Engineering.
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Affiliation(s)
- Daniela Patrício
- Institute of Biomedicine, Department of Medical Sciences, University of Aveiro, Aveiro, Portugal.,Department of Chemistry, CICECO - Aveiro Institute of Materials, University of Aveiro, Aveiro, Portugal
| | - Joana Santiago
- Institute of Biomedicine, Department of Medical Sciences, University of Aveiro, Aveiro, Portugal
| | - João F Mano
- Department of Chemistry, CICECO - Aveiro Institute of Materials, University of Aveiro, Aveiro, Portugal
| | - Margarida Fardilha
- Institute of Biomedicine, Department of Medical Sciences, University of Aveiro, Aveiro, Portugal
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10
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Greeson KW, Crow KMS, Edenfield RC, Easley CA. Inheritance of paternal lifestyles and exposures through sperm DNA methylation. Nat Rev Urol 2023:10.1038/s41585-022-00708-9. [PMID: 36653672 DOI: 10.1038/s41585-022-00708-9] [Citation(s) in RCA: 17] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/12/2022] [Indexed: 01/19/2023]
Abstract
Many different lifestyle factors and chemicals present in the environment are a threat to the reproductive tracts of humans. The potential for parental preconception exposure to alter gametes and for these alterations to be passed on to offspring and negatively affect embryo growth and development is of concern. The connection between maternal exposures and offspring health is a frequent focus in epidemiological studies, but paternal preconception exposures are much less frequently considered and are also very important determinants of offspring health. Several environmental and lifestyle factors in men have been found to alter sperm epigenetics, which can regulate gene expression during early embryonic development. Epigenetic information is thought to be a mechanism that evolved for organisms to pass on information about their lived experiences to offspring. DNA methylation is a well-studied epigenetic regulator that is sensitive to environmental exposures in somatic cells and sperm. The continuous production of sperm from spermatogonial stem cells throughout a man's adult life and the presence of spermatogonial stem cells outside of the blood-testis barrier makes them susceptible to environmental insults. Furthermore, altered sperm DNA methylation patterns can be maintained throughout development and ultimately result in impairments, which could predispose offspring to disease. Innovations in human stem cell-based spermatogenic models can be used to elucidate the paternal origins of health and disease.
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Affiliation(s)
- Katherine W Greeson
- Department of Environmental Health Science, College of Public Health, University of Georgia, Athens, GA, USA.,Regenerative Bioscience Center, University of Georgia, Athens, GA, USA
| | - Krista M S Crow
- Department of Environmental Health Science, College of Public Health, University of Georgia, Athens, GA, USA.,Regenerative Bioscience Center, University of Georgia, Athens, GA, USA
| | - R Clayton Edenfield
- Department of Environmental Health Science, College of Public Health, University of Georgia, Athens, GA, USA.,Regenerative Bioscience Center, University of Georgia, Athens, GA, USA
| | - Charles A Easley
- Department of Environmental Health Science, College of Public Health, University of Georgia, Athens, GA, USA. .,Regenerative Bioscience Center, University of Georgia, Athens, GA, USA.
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11
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Stanton JE, Grabrucker AM. The use of organoids in food research. Curr Opin Food Sci 2022. [DOI: 10.1016/j.cofs.2022.100977] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
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12
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Kouthouridis S, Robson E, Hartung A, Raha S, Zhang B. Se(XY) matters: the importance of incorporating sex in microphysiological models. Trends Biotechnol 2022; 40:1284-1298. [PMID: 35597689 DOI: 10.1016/j.tibtech.2022.04.005] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2022] [Revised: 04/06/2022] [Accepted: 04/09/2022] [Indexed: 01/21/2023]
Abstract
The development of microphysiological models is currently at the forefront of preclinical research. Although these 3D tissue models are being developed to mimic physiological organ function and diseases, which are often sexually dimorphic, sex is usually neglected as a biological variable. For decades, national research agencies have required government-funded clinical trials to include both male and female participants as a means of eliminating male bias. However, this is not the case in preclinical trials, which have been shown to favor male rodents in animal studies and male cell types in in vitro studies. In this Opinion, we highlight the importance of considering sex as a biological variable and outline five approaches for incorporating sex-specific features into current microphysiological models.
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Affiliation(s)
- Sonya Kouthouridis
- Department of Chemical Engineering, McMaster University, Hamilton, ON, L8S 4L8, Canada
| | - Eleanor Robson
- Department of Chemical Engineering, McMaster University, Hamilton, ON, L8S 4L8, Canada
| | - Alicia Hartung
- Department of Chemical Engineering, McMaster University, Hamilton, ON, L8S 4L8, Canada; School of Biomedical Engineering, McMaster University, Hamilton, ON, L8S 4L8, Canada
| | - Sandeep Raha
- Department of Pediatrics, McMaster University, Hamilton, ON, L8S 4L8, Canada; Graduate Program in Medical Sciences, McMaster University, Hamilton, ON, L8S 4L8, Canada.
| | - Boyang Zhang
- Department of Chemical Engineering, McMaster University, Hamilton, ON, L8S 4L8, Canada; School of Biomedical Engineering, McMaster University, Hamilton, ON, L8S 4L8, Canada.
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13
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Foster MJ, Patlewicz G, Shah I, Haggard DE, Judson RS, Paul Friedman K. Evaluating structure-based activity in a high-throughput assay for steroid biosynthesis. COMPUTATIONAL TOXICOLOGY (AMSTERDAM, NETHERLANDS) 2022; 24:1-23. [PMID: 37841081 PMCID: PMC10569244 DOI: 10.1016/j.comtox.2022.100245] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 10/17/2023]
Abstract
Data from a high-throughput human adrenocortical carcinoma assay (HT-H295R) for steroid hormone biosynthesis are available for >2000 chemicals in single concentration and 654 chemicals in multi-concentration (mc). Previously, a metric describing the effect size of a chemical on the biosynthesis of 11 hormones was derived using mc data referred to as the maximum mean Mahalanobis distance (maxmMd). However, mc HT-H295R assay data remain unavailable for many chemicals. This work leverages existing HT-H295R assay data by constructing structure-activity relationships to make predictions for data-poor chemicals, including: (1) identification of individual structural descriptors, known as ToxPrint chemotypes, associated with increased odds of affecting estrogen or androgen synthesis; (2) a random forest (RF) classifier using physicochemical property descriptors to predict HT-H295R maxmMd binary (positive or negative) outcomes; and, (3) a local approach to predict maxmMd binary outcomes using nearest neighbors (NNs) based on two types of chemical fingerprints (chemotype or Morgan). Individual chemotypes demonstrated high specificity (85-98%) for modulators of estrogen and androgen synthesis but with low sensitivity. The best RF model for maxmMd classification included 13 predicted physicochemical descriptors, yielding a balanced accuracy (BA) of 71% with only modest improvement when hundreds of structural features were added. The best two NN models for binary maxmMd prediction demonstrated BAs of 85 and 81% using chemotype and Morgan fingerprints, respectively. Using an external test set of 6302 chemicals (lacking HT-H295R data), 1241 were identified as putative estrogen and androgen modulators. Combined results across the three classification models (global RF model and two local NN models) predict that 1033 of the 6302 chemicals would be more likely to affect HT-H295R bioactivity. Together, these in silico approaches can efficiently prioritize thousands of untested chemicals for screening to further evaluate their effects on steroid biosynthesis.
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Affiliation(s)
- M J Foster
- Center for Computational Toxicology and Exposure, Office of Research and Development, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, 27711, USA
- National Student Services Contractor, Oak Ridge Associated Universities
| | - G Patlewicz
- Center for Computational Toxicology and Exposure, Office of Research and Development, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, 27711, USA
| | - I Shah
- Center for Computational Toxicology and Exposure, Office of Research and Development, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, 27711, USA
| | - D E Haggard
- Center for Computational Toxicology and Exposure, Office of Research and Development, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, 27711, USA
| | - R S Judson
- Center for Computational Toxicology and Exposure, Office of Research and Development, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, 27711, USA
| | - K Paul Friedman
- Center for Computational Toxicology and Exposure, Office of Research and Development, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, 27711, USA
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14
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Ma Y, Chen J, Li H, Xu F, Chong T, Wang Z, Zhang L. Immature rat testis sustained long-term development using an integrative model. Biol Res 2022; 55:30. [PMID: 36195947 PMCID: PMC9531454 DOI: 10.1186/s40659-022-00398-y] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/25/2022] [Accepted: 09/27/2022] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Xenotransplantation has been primarily performed using fresh donor tissue to study testicular development for about 20 years, and whether the cultured tissue would be a suitable donor is unclear. In this study, we combined testicular culture and xenotransplantation into an integrative model and explored whether immature testicular tissue would survive and continue to develop in this model. METHODS In the new integrative model group, the testes of neonatal rats on postnatal day 8 (PND 8) were cultured for 4 days ex vivo and then were transplanted under the dorsal skin of castrated nude mice. The xenografted testes were resected on the 57th day after xenotransplantation and the testes of rats in the control group were harvested on PND 69. The survival state of testicular tissue was evaluated from morphological and functional perspectives including H&E staining, immunohistochemical staining of 8-OH-dG, immunofluorescence staining, TUNEL assay, ultrastructural study, gene expression and protein analysis. RESULTS (a) We found that complete spermatogenesis was established in the testes in the new integrative model group. Compared with the control in the same stage, the seminiferous epithelium in some tubules was a bit thinner and there were vacuoles in part of the tubules. Immunofluorescence staining revealed some ACROSIN-positive spermatids were present in seminiferous tubule of xenografted testes. TUNEL detection showed apoptotic cells and most of them were germ cells in the new integrative model group. 8-OH-dG immunohistochemistry showed strongly positive-stained in the seminiferous epithelium after xenotransplantation in comparison with the control group; (b) Compared with the control group, the expressions of FOXA3, DAZL, GFRα1, BOLL, SYCP3, CDC25A, LDHC, CREM and MKI67 in the new integrative model group were significantly elevated (P < 0.05), indicating that the testicular tissue was in an active differentiated and proliferative state; (c) Antioxidant gene detection showed that the expression of Nrf2, Keap1, NQO1 and SOD1 in the new integrative model group was significantly higher than those in the control group (P < 0.05), and DNA methyltransferase gene detection showed that the expression of DNMT3B was significantly elevated as well (P < 0.05). CONCLUSION The new integrative model could maintain the viability of immature testicular tissue and sustain the long-term survival in vivo with complete spermatogenesis. However, testicular genes expression was altered, vacuolation and thin seminiferous epithelium were still apparent in this model, manifesting that oxidative damage may contribute to the testicular development lesion and it needs further study in order to optimize this model.
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Affiliation(s)
- Yubo Ma
- Department of Urology, The Second Affiliated Hospital of Xi'an Jiaotong University, No. 157 Xiwu Road, Xi'an, 710004, China
| | - Juan Chen
- Department of Urology, The Second Affiliated Hospital of Xi'an Jiaotong University, No. 157 Xiwu Road, Xi'an, 710004, China
| | - Hecheng Li
- Department of Urology, The Second Affiliated Hospital of Xi'an Jiaotong University, No. 157 Xiwu Road, Xi'an, 710004, China
| | - Fangshi Xu
- Department of Urology, The Second Affiliated Hospital of Xi'an Jiaotong University, No. 157 Xiwu Road, Xi'an, 710004, China
| | - Tie Chong
- Department of Urology, The Second Affiliated Hospital of Xi'an Jiaotong University, No. 157 Xiwu Road, Xi'an, 710004, China
| | - Ziming Wang
- Department of Urology, The Second Affiliated Hospital of Xi'an Jiaotong University, No. 157 Xiwu Road, Xi'an, 710004, China.
| | - Liandong Zhang
- Department of Urology, The Second Affiliated Hospital of Xi'an Jiaotong University, No. 157 Xiwu Road, Xi'an, 710004, China.
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15
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Tahmasbpour Marzouni E, Stern C, Henrik Sinclair A, Tucker EJ. Stem Cells and Organs-on-chips: New Promising Technologies for Human Infertility Treatment. Endocr Rev 2022; 43:878-906. [PMID: 34967858 DOI: 10.1210/endrev/bnab047] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/19/2021] [Indexed: 11/19/2022]
Abstract
Having biological children remains an unattainable dream for most couples with reproductive failure or gonadal dysgenesis. The combination of stem cells with gene editing technology and organ-on-a-chip models provides a unique opportunity for infertile patients with impaired gametogenesis caused by congenital disorders in sex development or cancer survivors. But how will these technologies overcome human infertility? This review discusses the regenerative mechanisms, applications, and advantages of different types of stem cells for restoring gametogenesis in infertile patients, as well as major challenges that must be overcome before clinical application. The importance and limitations of in vitro generation of gametes from patient-specific human-induced pluripotent stem cells (hiPSCs) will be discussed in the context of human reproduction. The potential role of organ-on-a-chip models that can direct differentiation of hiPSC-derived primordial germ cell-like cells to gametes and other reproductive organoids is also explored. These rapidly evolving technologies provide prospects for improving fertility to individuals and couples who experience reproductive failure.
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Affiliation(s)
- Eisa Tahmasbpour Marzouni
- Laboratory of Regenerative Medicine & Biomedical Innovations, Pasteur Institute of Iran, Tehran, Iran
| | - Catharyn Stern
- Royal Women's Hospital, Parkville and Melbourne IVF, Melbourne, Australia
| | - Andrew Henrik Sinclair
- Reproductive Development, Murdoch Children's Research Institute, Melbourne, Australia.,Department of Paediatrics, University of Melbourne, Melbourne, Australia
| | - Elena Jane Tucker
- Reproductive Development, Murdoch Children's Research Institute, Melbourne, Australia.,Department of Paediatrics, University of Melbourne, Melbourne, Australia
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16
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Jahanbani Y, Shafiee S, Davaran S, Roshangar L, Ahmadian E, Eftekhari A, Dolati S, Yousefi M. Stem cells technology as a platform for generating reproductive system organoids and treatment of infertility-related diseases. Cell Biol Int 2022; 46:512-522. [PMID: 34918417 DOI: 10.1002/cbin.11747] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2021] [Accepted: 12/14/2021] [Indexed: 02/06/2023]
Abstract
In recent years, stem cells have known as a helpful biological tool for the accurate diagnosis, treatment and recognition of diseases. Using stem cells as biomarkers have presented high potential in the early detection of many diseases. Another advancement in stem cell technology includes stem cell derived organoids model that could be a promising platform for diagnosis and modeling different diseases. Furthermore, therapeutic capabilities of stem cell therapy have increased hope in the face of different disability managements. All of these technologies are also widely used in reproductive related diseases especially in today's world that many couples encounter infertility problems. However, with the aid of numerous improvements in the treatment of infertility, over 80% of couples who dreamed of having children could now have children. Due to the fact that infertility has many negative effects on personal and social lives of young couples, many researchers have focused on the treatment of male and female reproductive system abnormalities with different types of stem cells, including embryonic stem cells, bone marrow mesenchymal stem cells (MSCs), and umbilical cord-derived MSCs. Also, design and formation of reproductive system organoids provide a fascinating window into disease modeling, drug screening, personalized therapy, and regeneration medicine. Utilizing these techniques to study, model and treat the infertility-related diseases has drawn attention of many scientists. This review explains different applications of stem cells in generating reproductive system organoids and stem cell-based therapies for male and female infertility related diseases treatment.
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Affiliation(s)
- Yalda Jahanbani
- Research Center for Pharmaceutical Nanotechnology, Tabriz University of Medical Sciences, Tabriz, Iran
- Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Samira Shafiee
- Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran
- Department of Medicinal Chemistry, Faculty of Pharmacy, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Soodabeh Davaran
- Department of Medicinal Chemistry, Faculty of Pharmacy, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Leila Roshangar
- Stem Cell Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Elham Ahmadian
- Kidney Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Aziz Eftekhari
- Stem Cell Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
- Russian Institute for Advanced Study, Moscow State Pedagogical University, Moscow, Russian Federation
| | - Sanam Dolati
- Physical Medicine and Rehabilitation Research Center, Aging Research Institute, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Mehdi Yousefi
- Department of Immunology, School of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran
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17
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Implications of testicular ACE2 and the renin-angiotensin system for SARS-CoV-2 on testis function. Nat Rev Urol 2022; 19:116-127. [PMID: 34837081 PMCID: PMC8622117 DOI: 10.1038/s41585-021-00542-5] [Citation(s) in RCA: 31] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 10/26/2021] [Indexed: 12/16/2022]
Abstract
Although many studies have focused on SARS-CoV-2 infection in the lungs, comparatively little is known about the potential effects of the virus on male fertility. SARS-CoV-2 infection of target cells requires the presence of furin, angiotensin-converting enzyme 2 (ACE2) receptors, and transmembrane protease serine 2 (TMPRSS2). Thus, cells in the body that express these proteins might be highly susceptible to viral entry and downstream effects. Currently, reports regarding the expression of the viral entry proteins in the testes are conflicting; however, other members of the SARS-CoV family of viruses - such as SARS-CoV - have been suspected to cause testicular dysfunction and/or orchitis. SARS-CoV-2, which displays many similarities to SARS-CoV, could potentially cause similar adverse effects. Commonalities between SARS family members, taken in combination with sparse reports of testicular discomfort and altered hormone levels in patients with SARS-CoV-2, might indicate possible testicular dysfunction. Thus, SARS-CoV-2 infection has the potential for effects on testis somatic and germline cells and experimental approaches might be required to help identify potential short-term and long-term effects of SARS-CoV-2 on male fertility.
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18
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Zarkower D, Murphy MW. DMRT1: An Ancient Sexual Regulator Required for Human Gonadogenesis. Sex Dev 2022; 16:112-125. [PMID: 34515237 PMCID: PMC8885888 DOI: 10.1159/000518272] [Citation(s) in RCA: 33] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2021] [Accepted: 05/25/2021] [Indexed: 01/03/2023] Open
Abstract
Transcriptional regulators related to the invertebrate sexual regulators doublesex and mab-3 occur throughout metazoans and control sex in most animal groups. Seven of these DMRT genes are found in mammals, and mouse genetics has shown that one, Dmrt1, plays a crucial role in testis differentiation, both in germ cells and somatic cells. Deletions and, more recently, point mutations affecting human DMRT1 have demonstrated that its heterozygosity is associated with 46,XY complete gonadal dysgenesis. Most of our detailed knowledge of DMRT1 function in the testis, the focus of this review, derives from mouse studies, which have revealed that DMRT1 is essential for male somatic and germ cell differentiation and maintenance of male somatic cell fate after differentiation. Moreover, ectopic DMRT1 can reprogram differentiated female granulosa cells into male Sertoli-like cells. The ability of DMRT1 to control sexual cell fate likely derives from at least 3 properties. First, DMRT1 functionally collaborates with another key male sex regulator, SOX9, and possibly other proteins to maintain and reprogram sexual cell fate. Second, and related, DMRT1 appears to function as a pioneer transcription factor, binding "closed" inaccessible chromatin and promoting its opening to allow binding by other regulators including SOX9. Third, DMRT1 binds DNA by a highly unusual form of interaction and can bind with different stoichiometries.
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Affiliation(s)
- David Zarkower
- Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, MN, USA
| | - Mark W. Murphy
- Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, MN, USA
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19
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Sekine K. Human Organoid and Supporting Technologies for Cancer and Toxicological Research. Front Genet 2021; 12:759366. [PMID: 34745227 PMCID: PMC8569236 DOI: 10.3389/fgene.2021.759366] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2021] [Accepted: 10/06/2021] [Indexed: 11/16/2022] Open
Abstract
Recent progress in the field of organoid-based cell culture systems has enabled the use of patient-derived cells in conditions that resemble those in cancer tissue, which are better than two-dimensional (2D) cultured cell lines. In particular, organoids allow human cancer cells to be handled in conditions that resemble those in cancer tissue, resulting in more efficient establishment of cells compared with 2D cultured cell lines, thus enabling the use of multiple patient-derived cells with cells from different genetic background, in keeping with the heterogeneity of the cells. One of the most valuable points of using organoids is that human cells from either healthy or cancerous tissue can be used. Using genome editing technology such as clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein, organoid genomes can be modified to, for example, cancer-prone genomes. The normal, cancer, or genome-modified organoids can be used to evaluate whether chemicals have genotoxic or non-genotoxic carcinogenic activity by evaluating the cancer incidence, cancer progression, and cancer metastasis. In this review, the organoid technology and the accompanying technologies were summarized and the advantages of organoid-based toxicology and its application to pancreatic cancer study were discussed.
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Affiliation(s)
- Keisuke Sekine
- Laboratory of Cancer Cell Systems, National Cancer Center Research Institute, Tokyo, Japan
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20
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Prasad M, Kumar R, Buragohain L, Kumari A, Ghosh M. Organoid Technology: A Reliable Developmental Biology Tool for Organ-Specific Nanotoxicity Evaluation. Front Cell Dev Biol 2021; 9:696668. [PMID: 34631696 PMCID: PMC8495170 DOI: 10.3389/fcell.2021.696668] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/17/2021] [Accepted: 08/13/2021] [Indexed: 12/14/2022] Open
Abstract
Engineered nanomaterials are bestowed with certain inherent physicochemical properties unlike their parent materials, rendering them suitable for the multifaceted needs of state-of-the-art biomedical, and pharmaceutical applications. The log-phase development of nano-science along with improved "bench to beside" conversion carries an enhanced probability of human exposure with numerous nanoparticles. Thus, toxicity assessment of these novel nanoscale materials holds a key to ensuring the safety aspects or else the global biome will certainly face a debacle. The toxicity may span from health hazards due to direct exposure to indirect means through food chain contamination or environmental pollution, even causing genotoxicity. Multiple ways of nanotoxicity evaluation include several in vitro and in vivo methods, with in vitro methods occupying the bulk of the "experimental space." The underlying reason may be multiple, but ethical constraints in in vivo animal experiments are a significant one. Two-dimensional (2D) monoculture is undoubtedly the most exploited in vitro method providing advantages in terms of cost-effectiveness, high throughput, and reproducibility. However, it often fails to mimic a tissue or organ which possesses a defined three-dimensional structure (3D) along with intercellular communication machinery. Instead, microtissues such as spheroids or organoids having a precise 3D architecture and proximate in vivo tissue-like behavior can provide a more realistic evaluation than 2D monocultures. Recent developments in microfluidics and bioreactor-based organoid synthesis have eased the difficulties to prosper nano-toxicological analysis in organoid models surpassing the obstacle of ethical issues. The present review will enlighten applications of organoids in nanotoxicological evaluation, their advantages, and prospects toward securing commonplace nano-interventions.
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Affiliation(s)
- Minakshi Prasad
- Department of Animal Biotechnology, Lala Lajpat Rai University of Veterinary and Animal Sciences, Hisar, India
| | - Rajesh Kumar
- Department of Veterinary Physiology and Biochemistry, Lala Lajpat Rai University of Veterinary and Animal Sciences, Hisar, India
| | - Lukumoni Buragohain
- Department of Animal Biotechnology, College of Veterinary Science, Assam Agricultural University, Guwahati, India
| | | | - Mayukh Ghosh
- Department of Veterinary Physiology and Biochemistry, RGSC, Banaras Hindu University, Varanasi, India
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21
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Abstract
In recent years, there has been an increased incidence in several of the most common reproductive disorders, including hypospadias and cryptorchidism in newborns, and testicular cancer and lower sperm quality in young adult males. In addition, the timing of puberty has also changed over time. Although the cause of these reproductive effects is a matter of intense debate, a link with the presence of ubiquitous compounds in the environment, or the exposure to specific groups of medications during foetal life, has been suggested. Results from epidemiological and experimental studies, as well as clinical observations in humans indicate that endocrine-disrupting chemicals may be associated with those disorders. In this review, we will summarize the results of epidemiological studies and experimental studies utilising human testicular cells or tissue. Due to increasing public interest and the recently published data, the main focus will be on the effects of prenatal exposure to mild analgesics.
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Affiliation(s)
- Jan-Bernd Stukenborg
- NORDFERTIL Research Lab Stockholm, Childhood Cancer Research Unit, Department of Women's and Children's Health, Karolinska Institutet, and Karolinska University Hospital, 17164 Solna, Sweden.
| | - Rod T Mitchell
- MRC Centre for Reproductive Health, The Queen's Medical Research Institute, The University of Edinburgh, and the Royal Hospital for Children and Young People, Edinburgh, UK.
| | - Olle Söder
- NORDFERTIL Research Lab Stockholm, Childhood Cancer Research Unit, Department of Women's and Children's Health, Karolinska Institutet, and Karolinska University Hospital, 17164 Solna, Sweden.
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22
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Cham TC, Ibtisham F, Fayaz MA, Honaramooz A. Generation of a Highly Biomimetic Organoid, Including Vasculature, Resembling the Native Immature Testis Tissue. Cells 2021; 10:cells10071696. [PMID: 34359871 PMCID: PMC8305979 DOI: 10.3390/cells10071696] [Citation(s) in RCA: 23] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/29/2021] [Revised: 06/27/2021] [Accepted: 07/04/2021] [Indexed: 12/25/2022] Open
Abstract
The creation of a testis organoid (artificial testis tissue) with sufficient resemblance to the complex form and function of the innate testis remains challenging, especially using non-rodent donor cells. Here, we report the generation of an organoid culture system with striking biomimicry of the native immature testis tissue, including vasculature. Using piglet testis cells as starting material, we optimized conditions for the formation of cell spheroids, followed by long-term culture in an air–liquid interface system. Both fresh and frozen-thawed cells were fully capable of self-reassembly into stable testis organoids consisting of tubular and interstitial compartments, with all major cell types and structural details expected in normal testis tissue. Surprisingly, our organoids also developed vascular structures; a phenomenon that has not been reported in any other culture system. In addition, germ cells do not decline over time, and Leydig cells release testosterone, hence providing a robust, tunable system for diverse basic and applied applications.
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Mollaki V. Ethical Challenges in Organoid Use. BIOTECH 2021; 10:biotech10030012. [PMID: 35822766 PMCID: PMC9245480 DOI: 10.3390/biotech10030012] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2021] [Revised: 05/31/2021] [Accepted: 06/23/2021] [Indexed: 12/12/2022] Open
Abstract
Organoids hold great promises for numerous applications in biomedicine and biotechnology. Despite its potential in science, organoid technology poses complex ethical challenges that may hinder any future benefits for patients and society. This study aims to analyze the multifaceted ethical issues raised by organoids and recommend measures that must be taken at various levels to ensure the ethical use and application of this technology. Organoid technology raises several serious ethics issues related to the source of stem cells for organoid creation, informed consent and privacy of cell donors, the moral and legal status of organoids, the potential acquisition of human “characteristics or qualities”, use of gene editing, creation of chimeras, organoid transplantation, commercialization and patentability, issues of equity in the resulting treatments, potential misuse and dual use issues and long-term storage in biobanks. Existing guidelines and regulatory frameworks that are applicable to organoids are also discussed. It is concluded that despite the serious ethical challenges posed by organoid use and biobanking, we have a moral obligation to support organoid research and ensure that we do not lose any of the potential benefits that organoids offer. In this direction, a four-step approach is recommended, which includes existing regulations and guidelines, special regulatory provisions that may be needed, public engagement and continuous monitoring of the rapid advancements in the field. This approach may help maximize the biomedical and social benefits of organoid technology and contribute to future governance models in organoid technology.
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Abstract
Organoids are 3-dimensional (3D) structures grown in vitro that emulate the cytoarchitecture and functions of true organs. Therefore, testicular organoids arise as an important model for research on male reproductive biology. These organoids can be generated from different sources of testicular cells, but most studies to date have used immature primary cells for this purpose. The complexity of the mammalian testicular cytoarchitecture and regulation poses a challenge for working with testicular organoids, because, ideally, these 3D models should mimic the organization observed in vivo. In this review, we explore the characteristics of the most important cell types present in the testicular organoid models reported to date and discuss how different factors influence the regulation of these cells inside the organoids and their outcomes. Factors such as the developmental or maturational stage of the Sertoli cells, for example, influence organoid generation and structure, which affect the use of these 3D models for research. Spermatogonial stem cells have been a focus recently, especially in regard to male fertility preservation. The regulation of the spermatogonial stem cell niche inside testicular organoids is discussed in the present review, as this research area may be positively affected by recent progress in organoid generation and tissue engineering. Therefore, the testicular organoid approach is a very promising model for male reproductive biology research, but more studies and improvements are necessary to achieve its full potential.
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Affiliation(s)
- Nathalia de Lima e Martins Lara
- Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, Alberta, Canada
| | - Sadman Sakib
- Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, Alberta, Canada
- Biochemistry and Molecular Biology, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada
| | - Ina Dobrinski
- Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, Alberta, Canada
- Biochemistry and Molecular Biology, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada
- Correspondence: Ina Dobrinski, DrMedVet, MVSc, PhD, Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, 404 HMRB, 3300 Hospital Dr NW, Calgary, Alberta T2N 4N1, Canada.
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Liu HC, Xie Y, Deng CH, Liu GH. Stem cell-based therapies for fertility preservation in males: Current status and future prospects. World J Stem Cells 2020; 12:1097-1112. [PMID: 33178394 PMCID: PMC7596443 DOI: 10.4252/wjsc.v12.i10.1097] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/26/2020] [Revised: 05/13/2020] [Accepted: 08/26/2020] [Indexed: 02/06/2023] Open
Abstract
With the decline in male fertility in recent years, strategies for male fertility preservation have received increasing attention. In this study, by reviewing current treatments and recent publications, we describe research progress in and the future directions of stem cell-based therapies for male fertility preservation, focusing on the use of spermatogonial stem cells (SSCs), SSC niches, SSC-based testicular organoids, other stem cell types such as mesenchymal stem cells, and stem cell-derived extracellular vesicles. In conclusion, a more comprehensive understanding of the germ cell microenvironment, stem cell-derived extracellular vesicles, and testicular organoids will play an important role in achieving male fertility preservation.
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Affiliation(s)
- Han-Chao Liu
- Department of Andrology, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou 510080, Guangdong Province, China
| | - Yun Xie
- Department of Andrology, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou 510080, Guangdong Province, China
| | - Chun-Hua Deng
- Department of Andrology, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou 510080, Guangdong Province, China
| | - Gui-Hua Liu
- Reproductive Medicine Research Center, The Sixth Affiliated Hospital of Sun Yat-sen University, Guangzhou 510655, Guangdong Province, China
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Edmonds ME, Woodruff TK. Testicular organoid formation is a property of immature somatic cells, which self-assemble and exhibit long-term hormone-responsive endocrine function. Biofabrication 2020; 12:045002. [PMID: 32492667 DOI: 10.1088/1758-5090/ab9907] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Testicular organoid models are tools to study testicular physiology, development, and spermatogenesis in vitro. However, few side-by-side comparisons of organoid generation method have been evaluated. Here, we directly tested whether the culture microenvironment is the prime determinant promoting testicular organoid self-assembly. Using Matrigel as a representative extracellular matrix (ECM), we compared multiple culture environments, 2D and 3D, ECM-free and ECM, for organoid self-assembly with immature murine testicular cells. De novo tissues were observed to self-assemble in all four culture environments tested within 72 h, however, these tissues only met requirements to be named organoids in 2D ECM and 3D ECM-free (3DF) culture methods. Based on these results, 3DF was selected for further study, and used to examine animal age as an independent variable. Organoid assembly was significantly delayed when using pubertal murine cells and entirely absent from adult murine and adult human cells. Organoid-conditioned medium and medium supplemented with 1% Matrigel did not improve organoid assembly in pubertal murine cells, but immature murine cells rescued the assembly of adult murine cells when cultured together as age-chimeric cell mixtures. In murine organoids cultured for 14 d, tubule-like structures exhibiting a highly biomimetic architecture were characterized, including some rare germ and spermatogonial stem cells. These structural organoids secreted high levels of testosterone and inhibin B over 12 weeks with preserved responsivity to gonadotropins. Collectively these studies, in which cellular self-assembly and organoid formation was achieved independent of the culture microenvironment, suggest that self-assembly is an innate property of immature testicular cells independent from, but capable of being promoted by, the culture environment. This study provides a template for studying testicular organoid self-assembly and endocrine function, and a platform for improving the engineering of functional testicular tissues.
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Affiliation(s)
- Maxwell E Edmonds
- Department of Obstetrics and Gynecology, Northwestern University Feinberg School of Medicine, Chicago, IL, United States of America
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