Empirical and physiological assessment of in vitro growth in the mermithid nematode Romanomermis culicivorax

1990 ◽  
Vol 68 (3) ◽  
pp. 511-516
Author(s):  
Roger Gordon ◽  
Marcel Cornect ◽  
Terry-Lynn Young ◽  
Kenneth T. Kean
Keyword(s):  
Lipid Content ◽  
Culture System ◽  
Culture Media ◽  
In Vitro Growth ◽  
Protein Levels ◽  
Physiological Assessment ◽  
Mermithid Nematode ◽  
Long Chain

In vitro growth of Romanomermis culicivorax preparasites was monitored in two culture media under differing conditions of crowding. Crowding was more important than the character of the medium itself in stimulating in vitro growth. The glycogen, protein, and lipid content of nematodes that had developed after 5 weeks under crowded conditions was significantly greater than that of uncrowded nematodes. The lipid content of such crowded nematodes was greater than that of nematodes after 4 days of in vivo development, although glycogen and protein levels were lower than the 4 day in vivo norm. Nematodes that had developed in vitro contained a higher proportion of long-chain and polyunsaturated fatty acids than did nematodes cultured in vivo. The significance of these findings is discussed in relation to the establishment of a continuous in vitro culture system for this nematode.

Embryonic development in culture of the marsupials Antechinus stuartii (Macleay) and Sminthopsis macroura (Spencer) during preimplantation stages

1993 ◽  
Vol 5 (4) ◽  
pp. 445 ◽  
Author(s):  
A Yousef ◽  
L Selwood
Keyword(s):  
Culture System ◽  
Fetal Calf Serum ◽  
Normal Development ◽  
Culture Media ◽  
Calf Serum ◽  
Human Amniotic Fluid ◽  
Antechinus Stuartii ◽  
Sminthopsis Macroura

Forty-nine blastocysts from 11 brown antechinus, Antechinus stuartii, and 96 blastocysts from 17 stripe-faced dunnarts, Sminthopsis macroura, were used to develop a culture system for embryos during preimplantation stages. Blastocysts of brown antechinus were collected on Days 6-9 for unilaminar stages, Days 16-21 for bilaminar stages and Days 20 and 21 for trilaminar stages. Blastocysts of stripe-faced dunnarts were collected on Day 6 for unilaminar stages, Days 6-8 for bilaminar stages and Day 8 for trilaminar stages. Culture media were Dulbecco's modified Eagle's medium (DMEM) with 4.5% glucose and Whittingham's T6 medium both of which were supplemented with 5, 10, 12.5 and 20% fetal calf serum (FCS). Antechinus serum (5%) and bovine serum albumin (0.1%, 0.2%) were also added to some media. Human amniotic fluid (HAF) and Monomed media were also tested. Blastocysts were cultured at 35 degrees C in 5% CO2 in air. DMEM + 10% FCS and HAF supported normal development for the longest periods and over the greatest range of stages. Developmental failure of blastocysts in vitro during expansion of the unilaminar blastocyst and formation of the bilaminar blastocyst suggests that these stages may be dependent on uterine signals. When cultured in DMEM + 10% FCS, the rate of development of bilaminar and trilaminar blastocysts into organogenesis was 4 h slower than in vivo in the stripe-faced dunnart and about 6 h slower than in vivo in the brown antechinus. Embryos of stripe-faced dunnarts were cultured to within 18 h of birth.

scholarly journals Osteogenic cells form mineralized particles, a few μm in size, in a 3D collagen gel culture

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e7889 ◽  
Author(s):  
Takanori Kihara ◽  
Chiya Umezu ◽  
Karin Sawada ◽  
Yukari Furutani
Keyword(s):  
Culture System ◽  
Culture Media ◽  
Collagen Gel ◽  
Matrix Vesicles ◽  
Osteogenic Cells ◽  
Collagen Gel Culture ◽  
3D Collagen ◽  
Gel Culture

Osteogenic cells form mineralized matrices in vitro, as well as in vivo. The formation and shape of the mineralized matrices are highly regulated by the cells. In vitro formation of mineralized matrices by osteogenic cells can be a model for in vivo osteogenesis. In this study, using a three-dimensional (3D) collagen gel culture system, we developed a new in vitro model for the formation of mineralized particles, a few µm in size, by the osteogenic cells. Human osteosarcoma (HOS) cells formed spherical mineralized matrices (about 12 µm) at approximately 7 days when cultured with β-glycerophosphate (β-GP)-containing culture media on 2D tissue culture plates. Alternately, when they were cultured in a 3D collagen gel containing β-GP, they formed mineralized particles with about 1.7 µm in the gel at approximately 3 days. Calcium precipitation in the gel was evaluated by measuring the gel turbidity. This type of mineralization of HOS cells, which formed mineralized particles inside the gel, was also observed in a peptide-based hydrogel culture. The mineralized particles were completely diminished by inhibiting the activity of Pit-1, phosphate cotransporter, of the HOS cells. When mouse osteoblast-like MC3T3-E1 cells, which form large and flat mineralized matrices in 2D osteogenic conditions at approximately 3 weeks of culture, were cultured in a 3D collagen gel, they also formed mineralized particles in the gel, similar to those in HOS cells, at approximately 18 days. Thus, osteogenic cells cultured in the 3D collagen gel form mineralized particles over a shorter period, and the mineralization could be easily determined by gel turbidity. This 3D gel culture system of osteogenic cells acts as a useful model for cells forming particle-type mineralized matrices, and we assume that the mineralized particles in the 3D hydrogel are calcospherulites, which are derived from matrix vesicles secreted by osteogenic cells.

The development of in vitro fertilized (ivf) cattle ova in an oviduct cell co-culture system

1992 ◽  
Vol 1992 ◽  
pp. 41-41
Author(s):  
C. McCaffrey ◽  
M.G. Diskin ◽  
K.H. Lu ◽  
J.M. Sreenan
Keyword(s):  
Culture System ◽  
Culture Media ◽  
Cell Stage ◽  
Conventional Culture ◽  
Large Numbers ◽  
Successful Production ◽  
Oviduct Cell ◽  
Developmental Block

There have been many recent reports of the successful production of large numbers of in vitro matured (IVM) and in vitro fertilised (IVF) cow ova. While a high proportion (80-90%) of oocytes undergo apparently normal maturation and fertilisation in vitro, only 15-20% develop in vitro to the morulae and blastocyst stages. Early cleavage stage in vivo matured and fertilised cow ova also encounter a developmental “block” at the 8-16-cell stage during in vitro culture in conventional culture media. However, this block has been readily overcome by co-culture involving the use of bovine oviducal cells, with up to 80% of 1-4-cell ova developing to morulae and blastocysts: The present study examined the development of IVM-IVF cattle ova removed at 24 or 48 hours from the insemination medium and cultured on cattle oviducal cells and also compared the development of IVM-IVF and in vivo produced ova in the same oviducal cell co-culture system.

90 LIPID CONTENT OF IN VIVO- AND IN VITRO-PRODUCED JERSEY AND HOLSTEIN CATTLE EMBRYOS AND THE EFFECT OF FORSKOLIN ON EMBRYO LIPID REDUCTION

2016 ◽  
Vol 28 (2) ◽  
pp. 174
Author(s):  
K. Rhodes-Long ◽  
L. F. Campos-Chillon ◽  
M. Barceló-Fimbres ◽  
J. L. Altermatt
Keyword(s):  
Fluorescence Intensity ◽  
Lipid Content ◽  
Lipid Accumulation ◽  
Culture Media ◽  
Nile Red ◽  
Holstein Cattle ◽  
Embryo Viability ◽  
Red Dye

Jersey embryos have been suggested to have higher lipid content and lower tolerance to cryopreservation. In addition, in vitro-produced (IVP) bovine embryos have darker cytoplasm as a consequence of higher lipid accumulation than in vivo-derived embryos, associated with impaired embryo quality and reduced cryotolerance. Forskolin is an adenylate cyclase activator that regulates cAMP levels in cells and has been shown to induce lipolysis in IVP embryos. We hypothesised that the lipid content of in vivo-produced and IVP Jersey embryos is higher than respective Holstein embryos and that forskolin would reduce lipid content of IVP embryos. The objectives of this experiment were (1) to analyse lipid content of in vivo and IVP Jersey and Holstein cattle embryos and (2) to evaluate the effect of forskolin added to IVP culture media. The factorial experimental design used two breeds (Holstein and Jersey) and three embryo production methods (in vivo, IVP, and IVP + forskolin). IVP embryos (n = 27 blastocysts) were collected from super-stimulated donors by routine procedures 7.5 days after AI. IVP embryos (n = 259 blastocysts) were produced by standard procedures; briefly, oocytes were aspirated from 2- to 8-mm follicles from slaughterhouse ovaries and matured for 24 h in SMM medium (BoviPro, MOFA Global, Verona, WI, USA). Matured oocytes were fertilized using semen from two different bulls for each breed, and embryos were cultured in BBH7 medium (BoviPro, MOFA Global) alone or with the addition of forskolin (10 µM) at Day 5 of culture at 38.5°C in 5% O2, 5% CO2, and 90% N2. The lipid content of embryos was quantified by staining Day 7 blastocysts with 1 μg mL–1 Nile red dye (580–596 nm), after which a digital photograph of the equatorial part of the embryo was taken at 40×, and fluorescence intensity (FI) was measured with Image Pro software. Data (Table 1) were analysed by ANOVA, and means were compared using Tukey’s HSD. Jersey and Holstein IVP embryos had higher lipid content than Holstein in vivo-produced embryos (P < 0.05), but were not different than Jersey in vivo-derived embryos (P > 0.1). Forskolin lowered the lipid content (P < 0.05) of both IVP Jersey and Holstein embryos and was not different (P > 0.1) than in vivo-produced embryos. Addition of forskolin to embryo culture media has the potential to lower embryo lipid accumulation and possibly improve embryo viability and cryotolerance of IVP embryos. Further studies including cryopreservation and transfer of IVP + forskolin embryos to recipients are necessary to corroborate the findings of the present study. Table 1.Fluorescence intensity of in vivo-produced and IVP Jersey and Holstein embryos

scholarly journals In vitro development of preimplantation porcine embryos using alginate hydrogels as a three-dimensional extracellular matrix

2014 ◽  
Vol 26 (7) ◽  
pp. 943 ◽  
Author(s):  
Catherine N. Sargus-Patino ◽  
Elane C. Wright ◽  
Sarah A. Plautz ◽  
Jeremy R. Miles ◽  
Jeff L. Vallet ◽  
...  
Keyword(s):  
Cell Survival ◽  
Culture System ◽  
Culture Media ◽  
Morphological Changes ◽  
Three Dimensional ◽  
In Vitro Development ◽  
Alginate Hydrogels ◽  
Vitro Development

Between Days 10 and 12 of gestation, porcine embryos undergo a dramatic morphological change, known as elongation, with a corresponding increase in oestrogen production that triggers maternal recognition of pregnancy. Elongation deficiencies contribute to embryonic loss, but exact mechanisms of elongation are poorly understood due to the lack of an effective in vitro culture system. Our objective was to use alginate hydrogels as three-dimensional scaffolds that can mechanically support the in vitro development of preimplantation porcine embryos. White cross-bred gilts were bred at oestrus (Day 0) to Duroc boars and embryos were recovered on Days 9, 10 or 11 of gestation. Spherical embryos were randomly assigned to be encapsulated within double-layered 0.7% alginate beads or remain as non-encapsulated controls (ENC and CONT treatment groups, respectively) and were cultured for 96 h. Every 24 h, half the medium was replaced with fresh medium and an image of each embryo was recorded. At the termination of culture, embryo images were used to assess morphological changes and cell survival. 17β-Oestradiol levels were measured in the removed media by radioimmunoassay. Real-time polymerase chain reaction was used to analyse steroidogenic transcript expression at 96 h in ENC and CONT embryos, as well as in vivo-developed control embryos (i.e. spherical, ovoid and tubular). Although no differences in cell survival were observed, 32% (P < 0.001) of the surviving ENC embryos underwent morphological changes characterised by tubal formation with subsequent flattening, whereas none of the CONT embryos exhibited morphological changes. Expression of steroidogenic transcripts STAR, CYP11A1 and CYP19A1 was greater (P < 0.07) in ENC embryos with morphological changes (ENC+) compared with CONT embryos and ENC embryos with no morphological changes (ENC–), and was more similar to expression of later-stage in vivo-developed controls. Furthermore, a time-dependent increase (P < 0.001) in 17β-oestradiol was observed in culture media from ENC+ compared with ENC– and CONT embryos. These results illustrate that preimplantation pig embryos encapsulated in alginate hydrogels can undergo morphological changes with increased expression of steroidogenic transcripts and oestrogen production, consistent with in vivo-developed embryos. This alginate culture system can serve as a tool for evaluating specific mechanisms of embryo elongation that could be targeted to improve pregnancy outcomes.

Effect of in Vivo Oxidized Cellulose on in Vitro Growth of Human Respiratory Mucosa and Sub-mucosa During Endoscopic Skull Base Approaches

2016 ◽  
Vol 77 (S 01) ◽  
Author(s):  
Ezequiel Goldschmidt ◽  
Jorge Rasmussen ◽  
Joseph Chabot ◽  
Monica Loressi ◽  
Marcelo Ielpi ◽  
...  
Keyword(s):  
Skull Base ◽  
Oxidized Cellulose ◽  
In Vitro Growth ◽  
Respiratory Mucosa

scholarly journals All-Trans Retinoic Acid Increases DRP1 Levels and Promotes Mitochondrial Fission

Cells ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 1202
Author(s):  
Bojjibabu Chidipi ◽  
Syed Islamuddin Shah ◽  
Michelle Reiser ◽  
Manasa Kanithi ◽  
Amanda Garces ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Mitochondrial Fission ◽  
Normal Function ◽  
Mitochondrial Network ◽  
Protein Levels ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Area And Perimeter

In the heart, mitochondrial homeostasis is critical for sustaining normal function and optimal responses to metabolic and environmental stressors. Mitochondrial fusion and fission are thought to be necessary for maintaining a robust population of mitochondria, and disruptions in mitochondrial fission and/or fusion can lead to cellular dysfunction. The dynamin-related protein (DRP1) is an important mediator of mitochondrial fission. In this study, we investigated the direct effects of the micronutrient retinoid all-trans retinoic acid (ATRA) on the mitochondrial structure in vivo and in vitro using Western blot, confocal, and transmission electron microscopy, as well as mitochondrial network quantification using stochastic modeling. Our results showed that ATRA increases DRP1 protein levels, increases the localization of DRP1 to mitochondria in isolated mitochondrial preparations. Our results also suggested that ATRA remodels the mitochondrial ultrastructure where the mitochondrial area and perimeter were decreased and the circularity was increased. Microscopically, mitochondrial network remodeling is driven by an increased rate of fission over fusion events in ATRA, as suggested by our numerical modeling. In conclusion, ATRA results in a pharmacologically mediated increase in the DRP1 protein. It also results in the modulation of cardiac mitochondria by promoting fission events, altering the mitochondrial network, and modifying the ultrastructure of mitochondria in the heart.

scholarly journals TRIM27 interacts with Iκbα to promote the growth of human renal cancer cells through regulating the NF-κB pathway

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Chengwu Xiao ◽  
Wei Zhang ◽  
Meimian Hua ◽  
Huan Chen ◽  
Bin Yang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Prognostic Indicator ◽  
The Cancer Genome Atlas ◽  
Mrna Levels ◽  
Loss Of Function ◽  
Protein Levels ◽  
Kaplan Meier ◽  
Cancer Genome Atlas

Abstract Background The tripartite motif (TRIM) family proteins exhibit oncogenic roles in various cancers. The roles of TRIM27, a member of the TRIM super family, in renal cell carcinoma (RCC) remained unexplored. In the current study, we aimed to investigate the clinical impact and roles of TRIM27 in the development of RCC. Methods The mRNA levels of TRIM27 and Kaplan–Meier survival of RCC were analyzed from The Cancer Genome Atlas database. Real-time PCR and Western blotting were used to measure the mRNA and protein levels of TRIM27 both in vivo and in vitro. siRNA and TRIM27 were exogenously overexpressed in RCC cell lines to manipulate TRIM27 expression. Results We discovered that TRIM27 was elevated in RCC patients, and the expression of TRIM27 was closely correlated with poor prognosis. The loss of function and gain of function results illustrated that TRIM27 promotes cell proliferation and inhibits apoptosis in RCC cell lines. Furthermore, TRIM27 expression was positively associated with NF-κB expression in patients with RCC. Blocking the activity of NF-κB attenuated the TRIM27-mediated enhancement of proliferation and inhibition of apoptosis. TRIM27 directly interacted with Iκbα, an inhibitor of NF-κB, to promote its ubiquitination, and the inhibitory effects of TRIM27 on Iκbα led to NF-κB activation. Conclusions Our results suggest that TRIM27 exhibits an oncogenic role in RCC by regulating NF-κB signaling. TRIM27 serves as a specific prognostic indicator for RCC, and strategies targeting the suppression of TRIM27 function may shed light on future therapeutic approaches.

scholarly journals Glucose Oligosaccharide and Long-Chain Glucomannan Feed Additives Induce Enhanced Activation of Intraepithelial NK Cells and Relative Abundance of Commensal Lactic Acid Bacteria in Broiler Chickens

2021 ◽  
Vol 8 (6) ◽  
pp. 110
Author(s):  
Nathalie Meijerink ◽  
Jean E. de Oliveira ◽  
Daphne A. van Haarlem ◽  
Guilherme Hosotani ◽  
David M. Lamot ◽  
...  
Keyword(s):  
Immune System ◽  
Nk Cells ◽  
Relative Abundance ◽  
Nk Cell ◽  
Feed Additives ◽  
Microbiota Composition ◽  
In Ovo ◽  
Long Chain

Restrictions on the use of antibiotics in the poultry industry stimulate the development of alternative nutritional solutions to maintain or improve poultry health. This requires more insight in the modulatory effects of feed additives on the immune system and microbiota composition. Compounds known to influence the innate immune system and microbiota composition were selected and screened in vitro, in ovo, and in vivo. Among all compounds, 57 enhanced NK cell activation, 56 increased phagocytosis, and 22 increased NO production of the macrophage cell line HD11 in vitro. Based on these results, availability and regulatory status, six compounds were selected for further analysis. None of these compounds showed negative effects on growth, hatchability, and feed conversion in in ovo and in vivo studies. Based on the most interesting numerical results and highest future potential feasibility, two compounds were analyzed further. Administration of glucose oligosaccharide and long-chain glucomannan in vivo both enhanced activation of intraepithelial NK cells and led to increased relative abundance of lactic acid bacteria (LAB) amongst ileum and ceca microbiota after seven days of supplementation. Positive correlations between NK cell subsets and activation, and relative abundance of LAB suggest the involvement of microbiota in the modulation of the function of intraepithelial NK cells. This study identifies glucose oligosaccharide and long-chain glucomannan supplementation as effective nutritional strategies to modulate the intestinal microbiota composition and strengthen the intraepithelial innate immune system.

scholarly journals Long noncoding RNA SNHG14 promotes hepatocellular carcinoma progression by regulating miR-876-5p/SSR2 axis

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Zhibin Liao ◽  
Hongwei Zhang ◽  
Chen Su ◽  
Furong Liu ◽  
Yachong Liu ◽  
...  
Keyword(s):  
Noncoding Rna ◽  
Animal Experiments ◽  
Luciferase Reporter ◽  
Western Blot Assay ◽  
Downstream Target ◽  
Protein Levels ◽  
Elevated Expression ◽  
Rna Immunoprecipitation

Abstract Background Aberrant expressions of long noncoding RNAs (lncRNAs) have been demonstrated to be related to the progress of HCC. The mechanisms that SNHG14 has participated in the development of HCC are obscure. Methods Quantitative real-time PCR (qRT-PCR) was used to measure the lncRNA, microRNA and mRNA expression level. Cell migration, invasion and proliferation ability were evaluated by transwell and CCK8 assays. The ceRNA regulatory mechanism of SNHG14 was evaluated by RNA immunoprecipitation (RIP) and dual luciferase reporter assay. Tumorigenesis mouse model was used to explore the roles of miR-876-5p in vivo. The protein levels of SSR2 were measured by western blot assay. Results In this study, we demonstrated that SNHG14 was highly expressed in HCC tissues, meanwhile, the elevated expression of SNHG14 predicted poor prognosis in patients with HCC. SNHG14 promoted proliferation and metastasis of HCC cells. We further revealed that SNHG14 functioned as a competing endogenous RNA (ceRNA) for miR-876-5p and that SSR2 was a downstream target of miR-876-5p in HCC. Transwell, CCK8 and animal experiments exhibited miR-876-5p inhibited HCC progression in vitro and in vivo. By conducting rescue experiments, we found the overexpression of SSR2 or knocking down the level of miR-876-5p could reverse the suppressive roles of SNHG14 depletion in HCC. Conclusion SNHG14 promotes HCC progress by acting as a sponge of miR-876-5p to regulate the expression of SSR2 in HCC.

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