METABOLIC CHANGES IN TESTICULAR CELLS FROM RATS AFTER LONG-TERM EXPOSURE TO 37 °C IN VIVO OR IN VITRO

1980 ◽  
Vol 85 (3) ◽  
pp. 471-479 ◽  
Author(s):  
F. F. G. ROMMERTS ◽  
F. H. DE JONG ◽  
J. A. GROOTEGOED ◽  
H. J. VAN DER MOLEN
Keyword(s):  
Sertoli Cells ◽  
Leydig Cells ◽  
Biochemical Properties ◽  
Effect Of Temperature ◽  
Seminiferous Tubules ◽  
Experimental Conditions ◽  
Testicular Cells ◽  
Old Rats

Biochemical properties of isolated Leydig cells, Sertoli cells and spermatocytes from rat testes have been investigated after in-vivo or in-vitro exposure of these cells to abdominal temperature (37 °C). The rate of production of testosterone and pregnenolone by isolated Leydig cells from cryptorchid and normal testes from mature rats was not different. Production of pregnenolone by mitochondria prepared from cryptorchid testes was 6·7 times higher than production by mitochondria from normal testes. Sertoli cells prepared from immature rats and incubated in vitro at 32 or 37 °C showed, on day 1 of the culture period, an initial twofold increase in the secretion of androgen-binding protein which was absent after 6 days in culture. In contrast, incorporation of [3H]leucine into secreted proteins was stimulated twofold on day 1 as well as by day 6 of culture. Secretion of oestradiol was increased 30-fold by day 6 when compared with the level found on day 1 when cells had been cultured at 37 °C and the increased secretion of oestradiol was maintained for approximately 2 days when the temperature of incubation was decreased to 32 °C Spermatocytes isolated from seminiferous tubules incubated for 20 h at 37 °C were active in the synthesis of RNA. No degeneration of these cells was observed in testes of 25-day-old rats 5 days after experimental cryptorchidism, whereas under similar conditions massive degeneration of spermatocytes was shown in the testes of mature rats. These results suggest that the effects of temperature on the different testicular cells greatly depend on the experimental conditions used to study the effect of temperature.

Applicability of Instability Index for In vitro Protein Stability Prediction

2019 ◽  
Vol 26 (5) ◽  
pp. 339-347 ◽  
Author(s):  
Dilani G. Gamage ◽  
Ajith Gunaratne ◽  
Gopal R. Periyannan ◽  
Timothy G. Russell
Keyword(s):  
Protein Stability ◽  
Caulobacter Crescentus ◽  
Effect Of Temperature ◽  
Instability Index ◽  
Dipeptide Composition ◽  
Experimental Conditions ◽  
The Stability ◽  
Vitro Protein

Background: The dipeptide composition-based Instability Index (II) is one of the protein primary structure-dependent methods available for in vivo protein stability predictions. As per this method, proteins with II value below 40 are stable proteins. Intracellular protein stability principles guided the original development of the II method. However, the use of the II method for in vitro protein stability predictions raises questions about the validity of applying the II method under experimental conditions that are different from the in vivo setting. Objective: The aim of this study is to experimentally test the validity of the use of II as an in vitro protein stability predictor. Methods: A representative protein CCM (CCM - Caulobacter crescentus metalloprotein) that rapidly degrades under in vitro conditions was used to probe the dipeptide sequence-dependent degradation properties of CCM by generating CCM mutants to represent stable and unstable II values. A comparative degradation analysis was carried out under in vitro conditions using wildtype CCM, CCM mutants and two other candidate proteins: metallo-β-lactamase L1 and α -S1- casein representing stable, borderline stable/unstable, and unstable proteins as per the II predictions. The effect of temperature and a protein stabilizing agent on CCM degradation was also tested. Results: Data support the dipeptide composition-dependent protein stability/instability in wt-CCM and mutants as predicted by the II method under in vitro conditions. However, the II failed to accurately represent the stability of other tested proteins. Data indicate the influence of protein environmental factors on the autoproteolysis of proteins. Conclusion: Broader application of the II method for the prediction of protein stability under in vitro conditions is questionable as the stability of the protein may be dependent not only on the intrinsic nature of the protein but also on the conditions of the protein milieu.

scholarly journals Only a small population of adult Sertoli cells actively proliferates in culture

Reproduction ◽  
2016 ◽  
Vol 152 (4) ◽  
pp. 271-281 ◽  
Author(s):  
Andrey Yu Kulibin ◽  
Ekaterina A Malolina
Keyword(s):  
Sertoli Cells ◽  
Collagen Matrix ◽  
Transitional Zone ◽  
Small Population ◽  
Seminiferous Tubules ◽  
Differentiated Cells ◽  
Low Levels ◽  
Two Populations

Adult mammalian Sertoli cells (SCs) have been considered to be quiescent terminal differentiated cells for many years, but recently, proliferation of adult SCs was demonstrated in vitro and in vivo. We further examined mouse SC behavior in culture and found that there are two populations of adult SCs. The first population is SCs from seminiferous tubules that hardly proliferate in vitro. The second population is small and consists of SCs with atypical nuclear morphology from the terminal segments of seminiferous tubules, a transitional zone (TZ). TZ SCs multiply in culture and form colonies, display mixture of mature and immature SC characteristics, and generate cord-like structures in a collagen matrix. The specific features of TZ SCs are ACTA2 expression in vitro and DMRT1 low levels in vivo and in vitro. Although the in vivo function of TZ SCs still remains unclear, this finding has significant implications for our understanding of SC differentiation and functioning in adult mammals.

Inhibin-like activity in Sertoli cell culture media and testicular homogenates from rats of various ages

1987 ◽  
Vol 113 (1) ◽  
pp. 103-110 ◽  
Author(s):  
A. M. Ultee-van Gessel ◽  
F. H. de Jong
Keyword(s):  
Sertoli Cells ◽  
Culture Media ◽  
Reciprocal Relationship ◽  
Pituitary Cells ◽  
In Vitro Experiments ◽  
Old Rats ◽  
Lh Secretion

ABSTRACT The influence of age on testicular inhibin in untreated, neonatally hemicastrated and prenatally irradiated rats was studied using in-vivo and in-vitro experiments. In testicular cytosols prepared from 1-, 7-, 14-, 21-, 42- and 63-day-old rats concentrations of testicular inhibin could be measured with an in-vitro bioassay method using dispersed pituitary cells. Preparations of testicular cytosols caused a dose-dependent suppression of pituitary FSH secretion, whereas no effects were found on LH secretion. Testicular content of inhibin increased gradually with age, while after 14 days of age a relatively large increase of peripheral FSH concentrations occurred in all experimental groups. Neonatal hemicastration or prenatal irradiation resulted in decreased inhibin content of the testis and increased plasma FSH levels. The production of inhibin activity by Sertoli cells obtained from 7-, 14-, 21-, 42- and 63-day-old normal rats was measured during a 24-h incubation period on the third day of culture. The inhibin production per 106 plated Sertoli cells decreased rapidly after 14 days of age and the lowest production of inhibin was found in Sertoli cells from rats of 63 days of age. After preincubation with ovine FSH significantly larger amounts of inhibin activity were detected in spent media from 21-day-old rat testes. In contrast, suppression of inhibin production was found after preculture in the presence of testosterone at most of the ages studied. These data from in-vivo and in-vitro experiments indicate that a reciprocal relationship exists between pituitary FSH secretion and inhibin production before the age of 21 days. This relationship supports the concept that inhibin is a physiologically important modulator of FSH secretion before puberty, while the role of the large amount of testicular inhibin present at the older ages remains to be determined. J. Endocr. (1987) 113, 103–110

scholarly journals Microfluidic bioprinting for the in vitro generation of novel biomimetic human testicular tissues

2021 ◽  
Author(s):  
Meghan Alice Robinson ◽  
Erin Bedford ◽  
Luke Witherspoon ◽  
Stephanie Willerth ◽  
Ryan Flannigan
Keyword(s):  
Cancer Survival ◽  
Fertility Restoration ◽  
Survival Rates ◽  
Testicular Tissue ◽  
Seminiferous Tubules ◽  
Vitro Fertilization ◽  
Testicular Cells ◽  
Human Spermatogenesis

Advances in cancer treatments have greatly improved pediatric cancer survival rates, leading to quality of life considerations and in particular fertility restoration. Accordingly, pre-pubertal patients have the option to cryopreserve testicular tissue for experimental restorative therapies, including in vitro spermatogenesis, wherein testicular tissue is engineered in vitro and spermatozoa are collected for in vitro fertilization (IVF). Current in vitro systems have been unable to reliably support the generation of spermatozoa from human testicular tissues, likely due to the inability for the dissociated testicular cells to recreate the native architecture of testicular tissue found in vivo. Recent advances in 3-D bioprinting can place cells into geometries at fine resolutions comparable to microarchitectures found in native tissues, and therefore hold promise as a tool for the development of a biomimetic in vitro system for human spermatogenesis. This study assessed the utility of bioprinting technology to recreate the precise architecture of testicular tissue and corresponding spermatogenesis for the first time. We printed testicular cell-laden hollow microtubules at similar resolutions to seminiferous tubules, and compared the results to testicular organoids. We show that the human testicular cells retain their viability and functionality post-printing, and illustrate an intrinsic ability to reorganize into their native cytoarchitecture. This study provides a proof of concept for the use of 3-D bioprinting technology as a tool to create biomimetic human testicular tissues.

The effect of prenatal treatment with busulfan on in vitro androgen production by testes from rats of various ages

1985 ◽  
Vol 63 (9) ◽  
pp. 1155-1158 ◽  
Author(s):  
Gwenderlyn F. Jansz ◽  
David K. Pomerantz
Keyword(s):  
Leydig Cells ◽  
Testicular Tissue ◽  
Developmental Changes ◽  
Seminiferous Tubules ◽  
Apparent Paradox ◽  
Old Rats ◽  
Effect Of Treatment ◽  
Hydroxy Steroid ◽  
Steroid Dehydrogenase

Treatment of rats with busulfan in utero severely depletes the germ cell population of the seminiferous tubules. These studies have examined the in vitro capacity of testicular tissue and Leydig cells from such testes to secrete androgens. Leydig cells were identified by staining for 3β-hydroxy steroid dehydrogenase. Rats were studied at several ages to identify any developmental changes in the androgen-secreting capacity of control and treated gonads. At 30 days of age, no effect of treatment on serum androgen was found. At 60 and 90 days of age, treatment caused decreased androgen and increased LH content of the serum. At 12, 30, 60, and 90 days of age, the amount of androgen secreted per milligram of testicular tissue in response to LH was higher in busulfan-treated rats. Leydig cells from 60- and 90-day-old rats which had received busulfan were also hyperresponsive to LH. It was concluded that Leydig cells from testes essentially devoid of germ cells were hyperresponsive to LH. Serum androgen levels were decreased yet androgen production per Leydig cell was increased. A possible explanation of this apparent paradox is that busulfan treatment resulted in decreased numbers of Leydig cells in the gonads.

scholarly journals Apoptotic spermatogenic cells can be energy sources for Sertoli cells

Reproduction ◽  
2009 ◽  
Vol 137 (3) ◽  
pp. 469-479 ◽  
Author(s):  
Weipeng Xiong ◽  
Haikun Wang ◽  
Hui Wu ◽  
Yongmei Chen ◽  
Daishu Han
Keyword(s):  
Sertoli Cells ◽  
Energy Sources ◽  
Seminiferous Tubules ◽  
Spermatogenic Cells ◽  
Atp Production ◽  
Lipid Formation ◽  
Residual Bodies ◽  
Induced Apoptosis

Apoptotic spermatogenic cells and residual bodies are phagocytosed and degraded by Sertoli cells during mammalian spermatogenesis. The meaning of this event remains to be clarified. In this report, we demonstrate that apoptotic spermatogenic cells and residual bodies can be used to produce ATP by Sertoli cells after phagocytosis of them. Sertoli cells produced the highest level of ATP compared with other testicular cells. Phagocytosis assayin vitroshowed that engulfment of apoptotic spermatogenic cells increases ATP production by Sertoli cells. The increased ATP production was detected in seminiferous tubules at the stages where phagocytosis occurs. Induced apoptosis of spermatogenic cellsin vivoincreased ATP production in seminiferous tubules. The augmentation of ATP production bothin vitroandin vivoassociated with the lipid formation in Sertoli cells after phagocytosis of apoptotic spermatogenic cells. The lipid β-oxidation was a predominant pathway to produce ATP in Sertoli cells. We conclude that after phagocytosis by Sertoli cells, apoptotic spermatogenic cells are degraded to form lipids that are then used to produce ATP. The results suggest that apoptotic spermatogenic cells can be energy sources for Sertoli cells that may define a novel meaning of spermatogenic cell death.

scholarly journals VCAM1-α4β1 integrin interaction mediates interstitial tissue reconstruction in 3-D re-aggregate culture of dissociated prepubertal mouse testicular cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Kazuko Abe ◽  
Shigeyuki Kon ◽  
Hiroki Kameyama ◽  
JiDong Zhang ◽  
Ken-ichirou Morohashi ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Leydig Cells ◽  
Interstitial Tissue ◽  
Testicular Cells ◽  
Aggregate Culture ◽  
Tissue Reconstruction ◽  
Blocking Antibodies ◽  
Cell To Cell Adhesion

AbstractRoles of interstitial tissue in morphogenesis of testicular structures remain less well understood. To analyze the roles of CD34+ cells in the reconstruction of interstitial tissue containing Leydig cells (LCs), and testicular structures, we used 3D-reaggregate culture of dissociated testicular cells from prepubertal mouse. After a week of culture, adult Leydig cells (ALCs) were preferentially incorporated within CD34+ cell-aggregates, but fetal LCs (FLCs) were not. Immunofluorescence studies showed that integrins α4, α9 and β1, and VCAM1, one of the ligands for integrins α4β1 and α9β1, are expressed mainly in CD34+ cells and ALCs, but not in FLCs. Addition of function-blocking antibodies against each integrin and VCAM1 to the culture disturbed the reconstruction of testicular structures. Antibodies against α4 and β1 integrins and VCAM1 robustly inhibited cell-to-cell adhesion between testicular cells and between CD34+ cells. Cell-adhesion assays indicated that CD34+ cells adhere to VCAM1 through the interaction with α4β1 integrin. Live cell imaging showed that CD34+ cells adhered around ALC-aggregates. CD34+ cells on the dish moved toward the aggregates, extending filopodia, and entered into them, which was disturbed by VCAM1 antibody. These results indicate that VCAM1-α4β1 integrin interaction plays pivotal roles in formation of testicular interstitial tissues in vitro and also in vivo.

Influence of thyroid hormone on androgen metabolism in peripuberal rat Sertoli cells

1994 ◽  
Vol 140 (3) ◽  
pp. 349-355 ◽  
Author(s):  
M L Panno ◽  
E Beraldi ◽  
V Pezzi ◽  
M Salerno ◽  
G De Luca ◽  
...  
Keyword(s):  
Body Weight ◽  
Thyroid Hormone ◽  
Sertoli Cells ◽  
Testosterone Metabolism ◽  
Androgen Metabolism ◽  
Functional Maturation ◽  
Severe Impairment ◽  
Old Rats

Abstract The aim of the present study was to investigate the influence of thyroid hormones on androgen metabolism in Sertoli cells isolated from 3- and 4- week-old rats. Hypothyroidism was induced by the oral administration of 0·025% methimazole (MMI) from birth until the rats were killed at 3 and 4 weeks of age. Half of the MMI-treated animals were injected i.p. with l-tri-iodothyronine (T3 3 μg/100 g body weight) during the last week before death. Sertoli cells from all groups were initially cultured under basal conditions for the first 24 h and subsequently in the presence of testosterone with or without T3 for an additional 24 h. Hypothyroidism was associated with severe impairment of body as well as testicular growth. Indeed, body and testicular weights were similar in 4-week-old hypothyroid animals to those in 3-week-old control rats. Testosterone metabolism in Sertoli cells isolated from 3- and 4-week-old hypothyroid rats was mainly expressed by the lowering of 5α-dihydrotestosterone + androstane 3α, 17β–diol and an enhanced formation of 5α-reduced steroids with poor androgenic properties (e.g. 5α–androstane, 3, 17α-dione (androstanedione), 5α–androstan, 3-ol-17-one (androsterone)). Treatment of the same group of animals with T3 in vivo and in vitro did not influence the pattern of 5α–reductase steroids substantially. The most striking finding in the Sertoli cells of 3-week-old hypothyroid rats was the dramatic enhancement of oestradiol formation which persisted to a lesser extent 1 week later. Oestradiol formation was greatly decreased by the addition of T3 in vivo and in vitro in hypothyroid animals. These results suggest that T3 might influence androgen metabolism during the functional maturation of Sertoli cells. Journal of Endocrinology (1994) 140, 349–355

FACTORS INFLUENCING THE UPTAKE OF [α-14C]AMINOISOBUTYRIC ACID BY RAT TESTES

1974 ◽  
Vol 60 (3) ◽  
pp. 463-471 ◽  
Author(s):  
O. IRUSTA ◽  
G. F. WASSERMANN
Keyword(s):  
Distribution Ratio ◽  
Follicle Stimulating Hormone ◽  
Aminoisobutyric Acid ◽  
Growing Rats ◽  
Testicular Cells ◽  
Factors Influencing ◽  
Old Rats ◽  
Cyclic Phosphate

SUMMARY The action of various agents on the distribution ratio of [α-14C]aminoisobutyric acid ([14C]AIB) in the testes of adult and growing rats was studied. A decrease in the distribution ratio of [14C]AIB was observed in the testes during maturation. Follicle-stimulating hormone (FSH) administered in vivo to 22-day-old rats or to mature hypophysectomized animals increased the distribution ratio of [14C]AIB in testicular cells. Dibutyryl adenosine 3′: 5′-cyclic phosphate augmented the [14C]AIB uptake into testes of intact or hypophysectomized mature animals. This nucleotide was ineffective in rats under 32 days of age. Testes from mature and immature rats preincubated in vitro with cycloheximide for 1 h showed a decrease in the distribution ratio of [14C]AIB. Cycloheximide also blocked the action of FSH on [14C]AIB uptake in the testes of 22-day-old rats.

scholarly journals Immunoexpression of Tyro 3 Family Receptors—Tyro 3, Axl, and Mer—and Their Ligand Gas6 in Postnatal Developing Mouse Testis

2005 ◽  
Vol 53 (11) ◽  
pp. 1355-1364 ◽  
Author(s):  
Huizhen Wang ◽  
Yongmei Chen ◽  
Yehua Ge ◽  
Pengpeng Ma ◽  
Quanhong Ma ◽  
...  
Keyword(s):  
Sertoli Cells ◽  
Leydig Cells ◽  
Cytoplasmic Membrane ◽  
Expression Patterns ◽  
Exact Expression ◽  
Seminiferous Tubules ◽  
Testicular Cells ◽  
Similar Expression Pattern ◽  
Weak Staining ◽  
Cell Processes

Tyro 3 family receptors contain three members—Tyro 3, Axl, and Mer—that are essential regulators of mammalian spermatogenesis. However, their exact expression patterns in testis are unclear. In this study, we examined the localizations of Tyro 3, Axl, Mer, and their ligand Gas6 in postnatal mouse testes by immunohistochemistry. All three members and their ligand were continuously expressed in different testicular cells during postnatal development. Tyro 3 was expressed only in Sertoli cells with a varied distribution during testis development. At day 3 postnatal, Tyro 3 was distributed in overall cytoplasmic membrane and cytoplasm of Sertoli cells. From day 14 to day 35 postnatal, Tyro 3 appeared on Sertoli cell processes toward the adlumenal compartment of seminiferous tubules. A stage-dependent Tyro 3 immunoexpression in Sertoli cells was shown by adulthood testis at day 56 postnatal with higher expression at stages I-VII and lower level at stages IX-XII. Axl showed a similar expression pattern to Tyro 3, except for some immunopositive Leydig cells detected in mature testis. In contrast, immunostaining of Mer was detected mainly in primitive spermatogonia and Leydig cells, whereas a relative weak signal was found in Sertoli cells. Gas6 was strongly expressed in Leydig cells, and a relative weak staining signal was seen in primitive spermatogonia and Sertoli cells. These immunoexpression patterns of Tyro 3 family receptors and ligand in testis provide a basis to further study their functions and mechanisms in regulating mammalian spermatogenesis.

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