Comparison of two culture methods during in vitro spermatogenesis of vitrified-warmed testis tissue: Organ culture vs. hanging drop culture

Cryobiology ◽  
2021 ◽  
Author(s):  
Tanushree Patra ◽  
Devendra Pathak ◽  
Mukesh Kumar Gupta
Keyword(s):  
Organ Culture ◽  
Hanging Drop ◽  
Culture Methods ◽  
In Vitro Spermatogenesis ◽  
Hanging Drop Culture ◽  
Testis Tissue

CIRCADIAN VARIATIONS IN ADRENAL SECRETION OF LEMMINGS, VOLES AND MICE

1970 ◽  
Vol 65 (4) ◽  
pp. 645-649 ◽  
Author(s):  
Richard V. Andrews
Keyword(s):  
Locomotor Activity ◽  
Organ Culture ◽  
Circadian Variation ◽  
The Arctic ◽  
Summer Solstice ◽  
Circadian Variations ◽  
Short Term ◽  
Culture Methods ◽  
Adrenal Secretion

ABSTRACT Daily variations in adrenal secretion by glands from arctic rodents were measured in vitro. Serial-sacrifice, short-term incubation studies yield similar results to those data obtained through organ-culture methods. Adrenal secretory rates display some desynchronization from locomotor activity during the summer solstice, although circadian variation persists during all seasons of the arctic year.

P–017 The maintenance of testicular architecture and germ cell in adult testis tissue under organ culture condition based on the gas-liquid interface method

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
M Komeya ◽  
H Odaka ◽  
T Matsumura ◽  
H Yamanaka ◽  
T Sato ◽  
...  
Keyword(s):  
Organ Culture ◽  
Germ Cell ◽  
Germ Cells ◽  
Culture Condition ◽  
Culture System ◽  
Liquid Interface ◽  
Adult Testis ◽  
In Vitro Spermatogenesis ◽  
Organ Culture System

Abstract Study question Can the gas-liquid interface organ culture system that achieved in vitro spermatogenesis in mice also support in vitro spermatogenesis in human adult testis? Summary answer Although the progression of spermatogenesis was not observed, germ cells were maintained without the degeneration of the architecture in both fresh and cryopreserved testicular tissues. What is known already Although the research on in vitro spermatogenesis have been conducted for 100 years, only the organ culture system using gas-liquid interface method achieved in vitro spermatogenesis in mice. It has not been verified whether this culture system can be applied to other mammals including humans and induce spermatogenesis. Study design, size, duration Testicular tissue was obtained from the transgender patients receiving sex reassignment surgery. Testicular specimens were either immediately processed for cultivation or cryopreserved, using a vitrification freezing protocol. Organ culture of testicular fragments was performed in three different media for a maximum period of 3 weeks to evaluate the short-term changes in the cultured tissues (viability, proliferation and maintenance of germ and somatic cells). Participants/materials, setting, methods Fresh and cryopreserved-thawed testis fragments (1–2 mm3) were cultured using the organ culture system in alpha-MEM with knock-out serum replacement (K group), alpha-MEM with lipid-rich BSA (A group) and DMEM with FBS (D group). Luteinizing hormone, follicle stimulating hormone and testosterone were supplemented. The number of germ cells (using DDX4), proliferative activity of germ cells (using EdU assay) and intratubular cell apoptosis (by TdT-mediated dUTP Nick End Labeling) were evaluated by immunohistochemical staining weekly. Main results and the role of chance The architecture of the seminiferous tubules was maintained until the second week of culture in both the fresh and the cryopreserved culture group. The number of DDX4-positive germ cells per seminiferous tubule in groups D, K, and A was 49 ± 24, 55 ± 21, 50 ± 26 cells/tubule in 1 day, 32 ± 13, 42 ± 7, 36 ± 21 cells/tubule in 1week, respectively. The numbers gradually decreased to 26 ± 8, 24 ± 6 and 27 ± 18 cells/tubule, in 2 weeks, respectively, with no difference among the groups. The number of intratubular EdU-positive cells of groups D, K, and A was 0.2 ± 0.2, 2.8 ± 2.1, 1.1 ± 0.8 cells/tubule at 1 day, 0.1 ± 0.2, 0.5 ± 0.6, 0.3 ± 0.6 cells/tubule at 1 week, respectively. The values were 0.01, 0.05, and 0.03 at 2 weeks. Thus, EdU-positive cells drastically decreased from the first week of culture. The number of DDX4-positive germ cells and the intratubular EdU-positive cells in the cryopreserved culture group was not different from that in the fresh culture group. Limitations, reasons for caution Current organ culture systems are incomplete, being unable to induce human in vitro spermatogenesis. Further research is needed to improve culture condition with the aim of producing fertile sperm of infertile adult male patients. Wider implications of the findings: Our organ culture system could maintain testis structure and germ cells. By using the testis tissues of the transgender patients, which are available with their consent, we will promote the investigation of the culture condition necessary for germ cell proliferation and differentiation. Trial registration number Grant-in-Aid for Scientific Research on Innovative Areas 18H05546, Grant-in-Aid for Young Scientists (A) 17H05098 and Takeda Science Foundation

174 In vitro spermatogenesis with a novel organ culture system using microfluidic technology

2015 ◽  
Vol 14 (2) ◽  
pp. e174-e174a
Author(s):  
M. Komeya ◽  
T. Yokonishi ◽  
T. Sato ◽  
K. Katagiri ◽  
M. Yao ◽  
...  
Keyword(s):  
Organ Culture ◽  
Culture System ◽  
In Vitro Spermatogenesis ◽  
Organ Culture System ◽  
Microfluidic Technology

scholarly journals Transcriptome analysis reveals inadequate spermatogenesis and immediate radical immune reactions during organ culture in vitro spermatogenesis

2020 ◽  
Vol 530 (4) ◽  
pp. 732-738 ◽  
Author(s):  
Takeru Abe ◽  
Hajime Nishimura ◽  
Takuya Sato ◽  
Harukazu Suzuki ◽  
Takehiko Ogawa ◽  
...  
Keyword(s):  
Organ Culture ◽  
Transcriptome Analysis ◽  
Culture In Vitro ◽  
Immune Reactions ◽  
In Vitro Spermatogenesis

In vitro spermatogenesis using bovine testis tissue culture techniques

2015 ◽  
Vol 12 (5) ◽  
pp. 314-323 ◽  
Author(s):  
Ki-Jung Kim ◽  
Byung-Gak Kim ◽  
Yong-Hee Kim ◽  
Yong-An Lee ◽  
Bang-Jin Kim ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Culture Techniques ◽  
In Vitro Spermatogenesis ◽  
Tissue Culture Techniques ◽  
Testis Tissue

scholarly journals A Dynamic Hanging-Drop System for Mesenchymal Stem Cell Culture

2020 ◽  
Vol 21 (12) ◽  
pp. 4298
Author(s):  
Shu-Wei Huang ◽  
Shian-Chiuan Tzeng ◽  
Jem-Kun Chen ◽  
Jui-Sheng Sun ◽  
Feng-Huei Lin
Keyword(s):  
Cell Culture ◽  
Spheroid Formation ◽  
Hanging Drop ◽  
Mesenchymal Stem Cell Culture ◽  
Physiological Conditions ◽  
The Past ◽  
Hanging Drop Culture ◽  
The Stability

There have been many microfluid technologies combined with hanging-drop for cell culture gotten developed in the past decade. A common problem within these devices is that the cell suspension introduced at the central inlet could cause a number of cells in each microwell to not regularize. Also, the instability of droplets during the spheroid formation remains an unsolved ordeal. In this study, we designed a microfluidic-based hanging-drop culture system with the design of taper-tube that can increase the stability of droplets while enhancing the rate of liquid exchange. A ring is surrounding the taper-tube. The ring can hold the cells to enable us to seed an adequate amount of cells before perfusion. Moreover, during the period of cell culture, the mechanical force around the cell is relatively low to prevent stem cells from differentiate and maintain the phenotype. As a result of our hanging system design, cells are designed to accumulate at the bottom of the droplet. This method enhances convenience for observation activities and analysis of experiments. Thus, this microfluid chip can be used as an in vitro platform representing in vivo physiological conditions, and can be useful in regenerative therapy.

In Vitro Spermatogenesis Using an Organ Culture Technique

2012 ◽  
pp. 479-488 ◽  
Author(s):  
Tetsuhiro Yokonishi ◽  
Takuya Sato ◽  
Kumiko Katagiri ◽  
Takehiko Ogawa
Keyword(s):  
Organ Culture ◽  
Culture Technique ◽  
In Vitro Spermatogenesis

IN VITRO METHODS FOR THE STUDY OF THE ADENOHYPOPHYSIAL FUNCTIONS TO SECRETE GONADOTROPHIN

1971 ◽  
Vol 68 (1_Suppl) ◽  
pp. S27-S40 ◽  
Author(s):  
T. Kobayashi ◽  
T. Kigawa ◽  
M. Mizuno ◽  
T. Watanabe
Keyword(s):  
Cell Culture ◽  
Organ Culture ◽  
Cultured Cells ◽  
Cell Sheet ◽  
Short Term ◽  
Hypothalamic Extract ◽  
Numerous Cell ◽  
Single Cell Culture ◽  
Almost All

ABSTRACT There are several in vitro methods to analyse the function of the adenohypophysis or the mechanisms of its regulation. The present paper deals with single cell culture, organ culture and short term incubation techniques by which the morphology and gonadotrophin-secreting function of the adenohypophysis were studied. In trypsin-dispersed cell culture, the adenohypophysial cells showed extensive propagation to form numerous cell colonies and finally develop into a confluent monolayer cell sheet covering completely the surface of culture vessels. Almost all of the cultured cells, however, became chromophobic, at least at the end of the first week of cultivation, when gonadotrophin was detectable neither in the culture medium nor in the cells themselves. After the addition of the hypothalamic extract, gonadotrophin became detectable again, and basophilic or PAS-positive granules also reappeared within the cells, suggesting that the gonadotrophs were stimulated by the extract to produce gonadotrophin. In organ culture and short term incubation, the incorporation of [3H] leucine into the adenohypophysial cells in relation to the addition of hypothalamic extract was examined. It was obvious that the ability to incorporate [3H] leucine into the gonadotrophs in vitro was highly dependent upon the presence of the hypothalamic extract.

Small Molecules Effective Against Liver and Blood Stage Malarial Infection

2019 ◽  
Vol 18 (23) ◽  
pp. 2008-2021 ◽  
Author(s):  
Snigdha Singh ◽  
Neha Sharma ◽  
Charu Upadhyay ◽  
Sumit Kumar ◽  
Brijesh Rathi ◽  
...  
Keyword(s):  
Drug Targets ◽  
Malaria Parasite ◽  
Blood Stage ◽  
Culture Methods ◽  
Liver Stage ◽  
Malarial Infection ◽  
Dual Stage ◽  
New Treatment ◽  
Global Threat

Malaria is a lethal disease causing devastating global impact by killing more than 8,00,000 individuals yearly. A noticeable decline in malaria related deaths can be attributed to the most reliable treatment, ACTs against P. falciparum. However, the cumulative resistance of the malaria parasite against ACTs is a global threat to control the disease and, therefore the new effective therapeutics are urgently needed, including new treatment approaches. Majority of the antimalarial drugs target BS malarial infection. Currently, scientists are eager to explore the drugs with potency against not only BS but other life stages such as sexual and asexual stages of the malaria parasite. Liver Stage is considered as one of the important drug targets as it always leads to BS and the infection can be cured at this stage before it enters into the Blood Stage. However, a limited number of compounds are reported effective against LS malaria infection probably due to scarcity of in vitro LS culture methods and clinical possibilities. This mini review covers a range of chemical compounds showing efficacy against BS and LS of the malaria parasite’s life cycle collectively (i.e. dual stage activity). These scaffolds targeting dual stages are essential for the eradication of malaria and to evade resistance.

scholarly journals Morpho-Physiological Testing of NaCl Sensitivity of Tobacco Plants Overexpressing Choline Oxidase Gene

Plants ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1102
Author(s):  
Galina N. Raldugina ◽  
Sergey V. Evsukov ◽  
Liliya R. Bogoutdinova ◽  
Alexander A. Gulevich ◽  
Ekaterina N. Baranova
Keyword(s):  
Choline Oxidase ◽  
Culture Methods ◽  
Gene Encoding ◽  
Oxidase Gene ◽  
Bacterial Enzyme ◽  
Transgenic Lines ◽  
High Regeneration ◽  
Whole Plants ◽  
Implicit Response

In this study the transgenic lines (TLs) of tobacco (Nicotianatabacum L.), which overexpress the heterologous gene encoding the bacterial enzyme choline oxidase were evaluated. The goal of our work is to study the effect of choline oxidase gene expression on the sensitivity of plant tissues to the action of NaCl. The regenerative capacity, rhizogenesis, the amount of photosynthetic pigments and osmotically active compounds (proline and glycine betaine) were assessed by in vitro cell culture methods using biochemical and morphological parameters. Transgenic lines with confirmed expression were characterized by high regeneration capacity from callus in the presence of 200 mmol NaCl, partial retention of viability at 400 mmol NaCl. These data correlated with the implicit response of regenerants and whole plants to the harmful effects of salinity. They turned out to be less sensitive to the presence of 200 mmol NaCl in the cultivation medium, in contrast to the WT plants.

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