In vitro proliferation and differentiation of mouse spermatogonial stem cells in decellularized human placenta matrix.

Utilizing natural scaffold production derived from extracellular matrix components presents a promising strategy for advancing in vitro spermatogenesis. In this study, we employed decellularized human placental tissue as a scaffold, upon which neonatal mouse spermatogonial cells (SCs) were cultured three-dimensional (3D) configuration. To assess cellular proliferation, we examined the expression of key markers (Id4 and Gfrα1) at both 1 and 14 days into the culture. Our quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis revealed a notable increase in Gfrα1 gene expression, with the 3D culture group exhibiting the highest levels. Furthermore, the relative frequency of Gfrα1-positive cells significantly rose from 38.1% in isolated SCs to 46.13% and 76.93% in the two-dimensional (2D) and 3D culture systems, respectively. Moving forward to days 14 and 35 of the culture period, we evaluated the expression of differentiating markers (Sycp3, acrosin, and Protamine 1). Sycp3 and Prm1 gene expression levels were upregulated in both 2D and 3D cultures, with the 3D group displaying the highest expression. Additionally, acrosin gene expression increased notably within the 3D culture. Notably, at the 35-day mark, the percentage of Prm1-positive cells in the 3D group (36.4%) significantly surpassed that in the 2D group (10.96%). This study suggests that the utilization of placental scaffolds holds significant promise as a bio-scaffold for enhancing mouse in vitro spermatogenesis.

Therapeutic potential of exosomes in spermatogenesis regulation and male infertility.

BACKGROUND: Spermatogenesis is a fundamental process crucial for male reproductive health and fertility. Exosomes, small membranous vesicles released by various cell types, have recently garnered attention for their role in intercellular communication. OBJECTIVE: This review aims to comprehensively explore the role of exosomes in regulating spermatogenesis, focusing on their involvement in testicular development and cell-to-cell communication. METHODS: A systematic examination of literature was conducted to gather relevant studies elucidating the biogenesis, composition, and functions of exosomes in the context of spermatogenesis. RESULTS: Exosomes play a pivotal role in orchestrating the complex signaling networks required for proper spermatogenesis. They facilitate the transfer of key regulatory molecules between different cell populations within the testes, including Sertoli cells, Leydig cells, and germ cells. CONCLUSION: The emerging understanding of exosome-mediated communication sheds light on novel mechanisms underlying spermatogenesis regulation. Further research in this area holds promise for insights into male reproductive health and potential therapeutic interventions.

Testicular niche repair after gonadotoxic treatments: Current knowledge and future directions.

The testicular niche, which includes the germ cells, somatic cells, and extracellular matrix, plays a crucial role in maintaining the proper functions of the testis. Gonadotoxic treatments, such as chemotherapy and radiation therapy, have significantly improved the survival rates of cancer patients but have also been shown to have adverse effects on the testicular microenvironment. Therefore, repairing the testicular niche after gonadotoxic treatments is essential to restore its function. In recent years, several approaches, such as stem cell transplantation, gene therapy, growth factor therapy, and pharmacological interventions have been proposed as potential therapeutic strategies to repair the testicular niche. This comprehensive review aims to provide an overview of the current understanding of testis damage and repair mechanisms. We will cover a range of topics, including the mechanism of gonadotoxic action, repair mechanisms, and treatment approaches. Overall, this review highlights the importance of repairing the testicular niche after gonadotoxic treatments and identifies potential avenues for future research to improve the outcomes for cancer survivors.

A framework for modeling an agronomic system's vulnerability to climate change with reflections from the Caspian coastal agro-ecological zone of Iran.

Assessing the vulnerability of different sectors to climate change has great importance in determining the appropriate adaptation measures to deal with climate change impacts on a river basin scale. In this research, using a framework for modeling the agronomic system vulnerability to climate change, vulnerability assessment under different scenarios was conducted for the Gorganrud River Basin located in the agro-ecological zone of the Caspian coastal plain of Iran. Considering exposure, susceptibility, and lack of resilience components, 12 indicators were chosen and quantified for both agronomic-environmental and socio-economic aspects. The SSM-iCrop2 model was used to simulate crop yield under current and climate change scenarios across the basin. The analysis indicates that in the current condition, the vulnerability level is different across the watersheds of the Gorganrud River Basin. By applying the climate change scenarios, agronomic system vulnerability would increase in the basin to some extent, particularly in Madarsu and Tilabad watersheds attributed with high vulnerability (0.63 and 0.61, respectively). This justifies the need to implement adaptation plans for encountering water shortage in the future. The analysis also suggests that the vulnerability of the agronomic system for adaptation scenarios characterized by less water consumption under climate change conditions is going to be slightly different from the vulnerability under the climate change scenarios. Due to an increase in agronomic system vulnerability under climate change scenarios, coupled with the fact that most watersheds (except Chehelchai, Nardin, and Narmab) are moderately vulnerable even under current conditions, policymakers and planners should promote crop and livelihood diversification programs aiming to prevent an increase in agronomic vulnerability.

Polycaprolactone/Testicular Extracellular Matrix/Graphene Oxide-Based Electrospun Tubular Scaffolds for Reproductive Medicine: Biomimetic Architecture of Seminiferous Tubules.

Numerous scaffolds are developed in the field of testicular bioengineering. However, effectively replicating the spatial characteristics of native tissue, poses a challenge in maintaining the requisite cellular arrangement essential for spermatogenesis. In order to mimic the structural properties of seminiferous tubules, the objective is to fabricate a biocompatible tubular scaffold. Following the decellularization process of the testicular tissue, validation of cellular remnants' elimination from the specimens is conducted using 4',6-diamidino-2-phenylindole staining, hematoxylin and eosin staining, and DNA content analysis. The presence of extracellular matrix (ECM) components is confirmed through Alcian blue, Orcein, and Masson's trichrome staining techniques. The electrospinning technique is employed to synthesize the scaffolds using polycaprolactone (PCL), extracted ECM, and varying concentrations of graphene oxide (GO) (0.5%, 1%, and 2%). Subsequently, comprehensive evaluations are performed to assess the properties of the synthetic scaffolds. These evaluations encompass Fourier-transform infrared spectroscopy, scanning electron microscopy imaging, scaffold degradation testing, mechanical behavior analysis, methylthiazolyldiphenyl-tetrazolium bromide assay, and in vivo biocompatibility assessment. The PCL/decellularized extracellular matrix with 0.5% GO formulation exhibits superior fiber morphology and enhanced mechanical properties, and outperforms other groups in terms of in vitro biocompatibility. Consequently, these scaffolds present a viable option for implementation in "in vitro spermatogenesis" procedures, holding promise for future sperm production from spermatogonial cells.

Ultrastructural study: in vitro and in vivo differentiation of mice spermatogonial stem cells.

Mouse testicular tissue is composed of seminiferous tubules and interstitial tissue. Mammalian spermatogenesis is divided into three stages: spermatocytogenesis (mitotic divisions) in which spermatogonial stem cells (SSCs) turn into spermatocytes, followed by two consecutive meiotic divisions in which spermatocytes form spermatids. Spermatids differentiate into spermatozoa during spermiogenesis. Various factors affect the process of spermatogenesis and the organization of cells in the testis. Any disorder in different stages of spermatogenesis will have negative effects on male fertility. The aim of the current study was to compare the in vitro and in vivo spermatogenesis processes before and after transplantation to azoospermic mice using ultrastructural techniques. In this study, mice were irradiated with single doses of 14 Gy 60Co radiation. SSCs isolated from neonatal mice were cultured in vitro for 1 week and were injected into the seminiferous tubule recipient's mice. Testicular cells of neonatal mice were cultured in the four groups on extracellular matrix-based 3D printing scaffolds. The transplanted testes (8 weeks after transplantation) and cultured testicular cells in vitro (after 3 weeks) were then processed for transmission electron microscopy studies. Our study's findings revealed that the morphology and ultrastructure of testicular cells after transplantation and in vitro culture are similar to those of in vivo spermatogenesis, indicating that spermatogenic cell nature is unaltered in vitro.

Enhancing differentiation of menstrual blood-derived stem cells into female germ cells using a bilayer amniotic membrane and nano-fibrous fibroin scaffold.

Three-dimensional nanofiber scaffolds offer a promising method for simulating in vivo conditions within the laboratory. This study aims to investigate the influence of a bilayer amniochorionic membrane/nanofibrous fibroin scaffold on the differentiation of human menstrual blood mesenchymal stromal/stem cells (MenSCs) into female germ cells. MenSCs were isolated and assigned to four culture groups: (i) MenSCs co-cultured with granulosa cells (GCs) using the scaffold (3D-T group), (ii) MenSCs using the scaffold alone (3D-C group), (iii) MenSCs co-cultured only with GCs (2D-T group), and (iv) MenSCs without co-culture or scaffold (2D-C group). Both MenSCs and GCs were independently cultured for two weeks before co-culturing was initiated. Flow cytometry was employed to characterize MenSCs based on positive markers (CD73, CD90, and CD105) and negative markers (CD45 and CD133). Additionally, flow cytometry and immunocytochemistry were used to characterize the GCs. Differentiated MenSCs were analyzed using real-time PCR and immunostaining. The real-time PCR results demonstrated significantly higher levels of VASA expression in the 3D-T group compared to the 3D-C, 2D-T, and 2D-C groups. Similarly, the SCP3 mRNA level in the 3D-T group was notably elevated compared to the 3D-C, 2D-T, and 2D-C groups. Moreover, the expression of GDF9 was significantly higher in the 3D-T group when compared to the 3D-C, 2D-T, and 2D-C groups. Immunostaining results revealed a lack of signal for VASA, SCP3, or GDF9 markers in the 2D-T group, while some cells in the 3D-T group exhibited positive staining for all these proteins. These findings suggest that the combination of a bilayer amniochorionic membrane/nanofibrous fibroin scaffold with co-culturing GCs facilitates the differentiation of MenSCs into female germ cells.

Exploring the influences of personal attitudes on the intention of continuing online grocery shopping after the COVID-19 pandemic.

The unprecedented COVID-19 pandemic has brought drastic changes in our daily activities. One of these essential activities is grocery shopping. In compliance with the recommended social distancing standards, many people have switched to online grocery shopping or curbside pickup to minimize possible contagion. Although the shift to online grocery shopping is substantial, it is not clear whether this change would last in the long term. This study examines the attributes and underlying attitudes that may influence individuals' future decisions on online grocery shopping. An online survey was conducted in May 2020 in South Florida to collect data for this study. The survey contained a comprehensive set of questions related to respondents' sociodemographic attributes, shopping and trip patterns, technology use, as well as attitudes toward telecommuting and online shopping. A structural equation model (SEM) was applied to examine the intervening effects of observed as well as latent attitude variables on the likelihood of online grocery shopping after the outbreak. The results indicated that those with more experience in using online grocery shopping platforms were more likely to continue purchasing their groceries online. Individuals with positive attitudes toward technology and online grocery shopping in terms of convenience, efficiency, usefulness, and easiness were more likely to adopt online grocery shopping in the future. On the other hand, pro- driving individuals were less likely to substitute online grocery shopping for in-store shopping. The results suggested that attitudinal factors could have substantial impacts on the propensity toward online grocery shopping.

Impacts of COVID-19 on Future Preferences Toward Telework.

This paper presents a study in capturing the impacts of the mandatory pandemic-induced telework practice on workers' perceptions of the benefits, challenges, and difficulties associated with telecommuting and how those might influence their preference for telework in the future. Data was collected through an online survey conducted in South Florida in May 2020. Survey data showed that telework indices (either measured through actual behavior or stated preference) before, during, and after the pandemic were heterogeneous across socio-economic, demographic, and attitudinal segments. Before the outbreak, males, full-time students, those with PhD degrees, and high-income people showed higher percentages of involvement in jobs with a telework option. They also had higher pro-technology, pro-online education, workaholic, and pro-telework attitudes. During the pandemic, professional/managerial/technical jobs as well as jobs with lower physical-proximity measures showed the highest telework frequency. In view of future telework preferences, our analysis showed that those who were more pro-telework, pro-technology, and showed less dislike of telework dislike preferred higher telework frequency. A structural equation model was developed to assess the impacts of different predictors on telework behavior before the pandemic and preferences after the pandemic. While telework frequency before the pandemic was highly affected by the pro-telework attitude, the after-pandemic preferences were influenced by several other attitudes such as dislike telework, enjoy interaction, workaholic, as well as productivity factors. This might confirm the assumption that the mandatory practice through the pandemic has provided employees more experiences with work-from-home arrangements, which could reshape decisions and expectations around telework adoption in the future.

Challenges, perceptions, and future preferences for post-secondary online education given experiences in the COVID-19 outbreak.

To gain a better understanding of online education status during and after the pandemic outbreak, this paper analyzed the data from a recent survey conducted in the state of Florida in May 2020. In particular, we focused on college students' perception of productivity changes, benefits, challenges, and their overall preference for the future of online education. Our initial exploratory analysis showed that in most cases, students were not fully satisfied with the quality of the online education, and the majority of them suffered a plummet in their productivities. Despite the challenges, around 61% believed that they would prefer more frequent participation in online programs in the future (compared to the normal conditions before the pandemic). A structural equation model was developed to identify and assess the factors that contribute to their productivity and future preferences. The results showed that lack of sufficient communication with other students/ instructor as well as lack of required technology infrastructure significantly reduced students' productivity. On the other hand, productivity was positively affected by perceived benefits such as flexibility and better time management. In addition, productivity played a mediating role for a number of socio-economic, demographic, and attitudinal attributes: including gender, income, technology attitudes, and home environment conflicts. Accordingly, females, high income groups, and those with home environment conflicts experienced lower productivity, which indirectly discouraged their preference for future online education. As expected, a latent pro-online education attitude increased both the productivity and the future online-education preference. Last but not the least, Gen-Xers were more likely to adopt online-education in the post pandemic conditions compared to their peers.

In vitro production of mouse morphological sperm in artificial testis bioengineered by 3D printing of extracellular matrix.

Since autologous stem cell transplantation is prone to cancer recurrence, in vitro sperm production is regarded a safer approach to fertility preservation. In this study, the spermatogenesis process on testicular tissue extracellular matrix (T-ECM)-derived printing structure was evaluated. Ram testicular tissue was decellularized using a hypertonic solution containing triton and the extracted ECM was used as a bio-ink to print an artificial testis. Following cell adhesion and viability examination, pre-meiotic and post-meiotic cells in the study groups (as testicular suspension and co-culture with Sertoli cells) were confirmed by real-time PCR, flow-cytometry and immunocytochemistry methods. Morphology of differentiated cells was evaluated using transmission electron microscopy (TEM), toluidine blue, Giemsa, and hematoxylin and eosin (H&E) staining. The functionality of Leydig and Sertoli cells was determined by their ability for hormone secretion. The decellularization of testicular tissue fragments was successful and had efficiently removed the cellular debris and preserved the ECM compounds. High cell viability, colonization, and increased expression of pre-meiotic markers in cultured testicular cells on T-ECM-enriched scaffolds confirmed their proliferation. Furthermore, the inoculation of neonatal mouse testicular cells onto T-ECM-enriched scaffolds resulted in the generation of sperm. Morphology evaluation showed that the structure of these cells was quite similar to mature sperm with a specialized tail structure. The hormonal analysis also confirmed production and secretion of testosterone and inhibin B by Leydig and Sertoli cells. T-ECM printed artificial testis is a future milestone that promises for enhancing germ cell maintenance and differentiation, toxicology studies, and fertility restoration to pave the way for new human infertility treatments in the future.

In vitro elimination of EL4 cancer cells from spermatogonia stem cells by miRNA-143- and 206-loaded folic acid-conjugated PLGA nanoparticles.

Aim: MiRNA's-143 and -206 are powerful apoptotic regulators in cancer cells. This study aimed to use miRNA-143- and 206-loaded poly(lactic-co-glycolic) acid (PLGA) nanoparticles conjugated with folic acid to induce apoptosis in the EL4 cancer cells. Materials & methods: The therapy was conducted in six groups: treatment with both miRNAs simultaneously (mixed miRNAs), miRNA-206 treatment, miRNA-143 treatment, blank PLGA, blank polyethylenimine (PEI) and complex PEI-miRNAs. Results: In terms of viability, in mixed miRNAs no synergistic effect was observed on EL4 cell elimination. However, in the single miRNA-206 group, a stronger apoptotic effect was observed than the mixed miRNAs group and single miRNA-143 group alone. Conclusion: MiRNAs' apoptotic induction effects in cancer cells were found to be remarkable.

Artificial testis: a testicular tissue extracellular matrix as a potential bio-ink for 3D printing.

Testicular scaffolds may be an option for fertility preservation. The aim was to develop various procedures for the decellularization of testicular tissue and to design a bio-ink to construct a bioartificial testis. Ram testicular tissue fragments were decellularized using NaCl buffer, NaCl buffer-Triton, SDS and SDS-Triton. The removal of the cells from the tissues was confirmed by DAPI and H & E staining, as well as the evaluation of the DNA content. Alcian blue, Orcein and Masson's trichrome staining methods were also used to confirm that T-ECM was preserved intact. Then, the optimal decellularization protocol was selected to determine the parameters of the bio-ink and printing of the scaffold. The extracted T-ECM was used to print the hydrogel scaffold in combination with alginate-gelatin. The printability, morphological, mechanical and biological properties of the printed hydrogels were characterized. Decellularization of testicular tissue fragments using the NaCl buffer-Triton protocol was significantly more efficient than other decellularization methods in removing the cellular debris and preserving the T-ECM compounds. The 3D printed scaffold with 5% T-ECM showed a uniform surface morphology with high cell attachment and cyto-biocompatibility properties for spermatogonia stem cells in vitro and in vivo compared to other groups. It is concluded that T-ECM can be used as a biomimetic material to make an artificial testis with possible in vitro sperm production.

Severity analysis for large truck rollover crashes using a random parameter ordered logit model.

Large truck rollover crashes present significant financial, industrial, and social impacts. This paper presents an effort to investigate the contributing factors to large truck rollover crashes. Specific focus was placed on exploring the role of heterogeneity and the potential sources of heterogeneity regarding their impacts on injury-severity outcomes. The data used in this study contained large truck rollover crashes that occurred between 2007 and 2016 in the state of Florida. A random parameter ordered logit (RPOL) model was applied. Various driver, vehicle, roadway, and crash attributes were explored as potential predictors in the model. Their impacts were examined for the presence of heterogeneity. Interaction effects were then added to the random variables in order to detect potential sources of heterogeneity. Model results showed that the impacts of lighting conditions and driving speed had significant variation across observations, and this variation could be attributed to driver actions and driver conditions at the time of the crash, as well as driver vision obstruction. Findings from this study shed light on the direction, magnitude, and randomness of the factors that contribute to large truck rollover crashes. Findings associated with heterogeneity could help develop more effective and targeted countermeasures to improve freight safety. Driver education programs could be planned more efficiently, and advisory and warning signs could be designed in a more insightful manner by taking into account specific roadway attributes, such as sandy surfaces, downhill, curved alignment, unpaved shoulders, and lighting conditions.

Future of Spermatogonial Stem Cell Culture: Application of Nanofiber Scaffolds.

BACKGROUND: Spermatogonial stem cells (SSCs) are unique in mammals because they can transmit genetic information from generation to generation and it is of significant importance. In testes, Sertoli cells, peritubular myoid cells, Leydig cells and other interstitial cells contribute to the spermatogonial stem cell "niche". So, creation of niche in an in vitro condition that mimics the in vivo environment is essential to maintain functional characteristic of SSCs. OBJECTIVE: In this review, we describe the impact of nanofiber scaffolds on the culture of SSCs derived from human-to-mouse. RESULTS: Nanofiber Matrices mimic the architecture and size scale of the natural extracellular matrix (ECM). The scaffold provides more three-dimensional (3D), biomimicking and topographical signals to the cells and results in a more physiologically relevant cellular phenotype. Several investigators use different nanofiber scaffold-like carbon nanotubes (CNTs) scaffold, poly-L-lactic acid (PLLA) nanofiber scaffold, 3D soft agar culture system, human serum albumin (HSA)/tri calcium phosphate nanoparticles (TCPNPs) and electrospun polyamide nanofiber for proliferation and maintenance of self-renewal activity of the SSCs. CONCLUSION: Application of nanofiber scaffolds for in vitro culture of the SSCs may produce spermatogonial stem cells that can be used in regenerative medicine, tissue engineering, assisted reproductive technology and in the treatment of infertility in pre-pubertal cancer patients.