scholarly journals Effect of spermine synthase deficiency on polyamine biosynthesis and content in mice and embryonic fibroblasts, and the sensitivity of fibroblasts to 1,3-bis-(2-chloroethyl)-N-nitrosourea

2000 ◽  
Vol 351 (2) ◽  
pp. 439-447 ◽  
Author(s):  
Caroline A. MACKINTOSH ◽  
Anthony E. PEGG
Keyword(s):  
The Body ◽  
Normal Male ◽  
Male Mice ◽  
Embryonic Fibroblasts ◽  
Primary Fibroblast ◽  
Polyamine Biosynthesis ◽  
Spermine Synthase ◽  
Fibroblast Cultures ◽  
Phex Gene ◽  
Weaning Age

Mutant Gy male mice, which have previously been described as having disruption of the phosphate-regulating Phex gene and a spermine synthase gene [Meyer, Henley, Meyer, Morgan, McDonald, Mills and Price (1998) Genomics, 48, 289–295; Lorenz, Francis, Gempel, Böddrich, Josten, Schmahl and Schmidt (1998) Hum. Mol. Genet. 7, 541–547], as well as mutant Hyp male mice, which have disruption of the Phex gene only, were examined along with their respective normal male littermates. Biochemical analyses of extracts of brains, hearts and livers of 5-week-old mice showed that Gy males lacked any significant spermine synthase activity as well as spermine content. Organs of Gy males had a higher spermidine content. This was caused not only by the lack of conversion of spermidine into spermine, but also because of compensatory increases in the activities of other polyamine biosynthetic enzymes. Gy males were half the body weight of their normal male littermates at weaning age. Hyp males, however, were no different in size when compared with their controls. High mortality of Gy males occurs by weaning age and this mortality was shown to be largely post-natal. Embryonic fibroblasts were isolated from Gy males and their normal male littermates and were similarly shown to lack any significant spermine synthase activity as well as spermine content. The lack of spermine, however, had no significant effect on the growth of immortalized fibroblasts or of primary fibroblast cultures. Similarly, there was no difference in the time of senescence of primary fibroblast cultures from Gy males compared with cultures derived from normal male littermates. However, the lack of spermine did increase the sensitivity of immortalized fibroblasts to killing by the chloroethylating agent 1,3-bis(2-chloroethyl)-N-nitrosourea. Therefore both the Gy male mice and derived embryonic fibroblasts provide valuable models to study the importance of spermine and spermine synthase, without the use of inhibitors which may have additional side effects.

Effect of polyamine depletion on caspase activation: a study with spermine synthase-deficient cells

2001 ◽  
Vol 355 (1) ◽  
pp. 199-206 ◽  
Author(s):  
Claudio STEFANELLI ◽  
Carla PIGNATTI ◽  
Benedetta TANTINI ◽  
Monia FATTORI ◽  
Ivana STANIC ◽  
...  
Keyword(s):  
Uv Irradiation ◽  
Normal Male ◽  
Caspase Activity ◽  
Caspase Activation ◽  
Embryonic Fibroblasts ◽  
Polyamine Synthesis ◽  
Spermine Synthase ◽  
Cell Processes ◽  
Mutant Male

Activation of the caspase proteases represents a central point in apoptosis. The requirement for spermine for the processes leading to caspase activation has been studied in transformed embryonic fibroblasts obtained from gyro (Gy) mutant male mice. These cells lack spermine synthase activity and thus provide a valuable model to study the role of spermine in cell processes. Gy fibroblasts do not contain spermine and have a higher spermidine content. However, when compared with fibroblasts obtained from normal male littermates (N cells), Gy fibroblasts were observed to grow normally. The lack of spermine did not affect the expression of Bcl-2, and caspases 3 and 9 were activated by etoposide in both N and Gy cells, indicating that spermine is dispensable for caspase activation. Spermine deficiency did not significantly influence caspase activity in cells treated with etoposide, cycloheximide or staurosporine, but sensitized the cells to UV irradiation, which triggered significantly higher caspase activity in Gy cells compared with N cells. α-Difluoromethylornithine (DFMO), an inhibitor of polyamine synthesis that is able to deplete cells of putrescine and spermidine, but usually does not influence spermine content, was able to produce a more complete polyamine depletion in Gy cells. This depletion, which included spermine deficiency, dramatically increased caspase activation and cell death in Gy fibroblasts exposed to UV irradiation. On the other hand, in either N or Gy cells, DFMO treatment did not influence caspase activity triggered by staurosporine, but inhibited it when the inducers were cycloheximide or etoposide. In Gy cells depleted of polyamines by DFMO, polyamine replenishment with either spermidine or spermine was sufficient to restore caspase activity induced by etoposide, indicating that, in this model, polyamines have an interchangeable role in supporting caspase activation. Therefore, spermine is not required for such activation, and the effect and specificity of polyamine depletion on caspase activity may be very different, depending on the role of polyamines in the specific death pathways engaged by different stimuli. Some inducers of apoptosis, for example etoposide, absolutely require polyamines for caspase activation, yet the lack of polyamines, particularly spermine, strongly increases caspase activation when induced by UV irradiation.

Abstract 620: Development Of A New Mouse Model Exhibiting A Specific Deficiency Of Prorenin Renin Receptor In Adipocytes.

Hypertension ◽  
2014 ◽  
Vol 64 (suppl_1) ◽  
Author(s):  
Frederique B Yiannikouris ◽  
Genevieve Nguyen
Keyword(s):  
Blood Pressure ◽  
Adipose Tissue ◽  
Mouse Model ◽  
Preliminary Data ◽  
The Body ◽  
Chow Diet ◽  
Slight Reduction ◽  
Male Mice ◽  
Exon 2 ◽  
Adipose Tissue Stromal Cells

Objectives: Recent studies demonstrated that the prorenin renin receptor (PRR) is present in adipose tissue. In adipose tissue stromal cells, PRR has the ability to bind renin and prorenin and contributes to the generation of angiotensinI (AngI). However, the contribution of adipocyte PRR to the generation of the vasoactive peptide, AngII and therefore to the regulation of blood pressure in physiological condition is unknown. The purpose of this study was to develop and characterize a new mouse model with adipocyte-specific PRR deficiency and define the role of adipose PRR in normal physiology. Methods and results: Female mice with 2 loxP sites flanking exon 2 of the PRR gene (floxed alleles, PRRfl/fl) were bred with aP2-Cre or with Adi-Cre male mice. Since PRR is located in the X chromosome, the male mice generated from the breeding were homozygotes for the deletion (PRRaP2 and PRRAdi). From the breeding, 5 PRRfl/fl, 2 PRRaP2 and 5 PRRAdi male mice were generated suggesting that the deletion of PRR in adipocyte was not lethal. Mice were fed on chow diet during 20 weeks. The body weight, the fat, lean mass and the blood pressure were quantified. Preliminary data suggest that the body weights (BW) were slightly decreased in PRRaP2 and PRRAdi compared to PRRfl/fl (PRRfl/fl: 29±1g; PRRaP2: 25±5g; PRRAdi: 28±1g). The slight reduction in BW was attributed to a reduction in fat mass (PRRfl/fl: 4.8±0.9g; PRRaP2: 3.8±1.8g; PRRAdi: 1.9±0.4g). Blood pressure was measured by plethysmography and by radiotelemetry. Preliminary data demonstrated that under physiological conditions, the SBP was not changed in PRRaP2 male mice compared to PRRfl/fl mice (plethysmography: PRRfl/fl: 108±1 mmHg; PRRaP2: 99±7 mmHg; radiotelemetry: PRRfl/fl: 129±2 mmHg; PRRaP2: 128±6 mmHg). The SBP of PRRAdi is currently under investigation. Conclusions: These results demonstrate the viability of mice with specific adipocyte deficiency of PRR. Future studies will define the effects of adipocyte PRR deficiency on obesity-induced hypertension.

IgG modulation in male mice with reproductive failure after prenatal inflammation

Reproduction ◽  
2021 ◽  
Author(s):  
Marina Izvolskaia ◽  
Vasilina Ignatiuk ◽  
Ayshat Ismailova ◽  
Viktoria Sharova ◽  
Liudmila Zakharova
Keyword(s):  
Positive Impact ◽  
Brain Structures ◽  
Peritoneal Macrophages ◽  
The Body ◽  
Male Rats ◽  
Male Mice ◽  
Prenatal Inflammation ◽  
Exposure In Vivo

Sexual performance in adult male rats is highly sensitive to prenatal stress which can affect the functionality of the reproductive system and various brain structures involved in modulating sexual behavior. The immunomodulatory effect of mouse IgG on reproductive maturity in male offspring after LPS exposure in vivo and in vitro was studied. Prenatal IgG injection (20 µg / mouse) had a positive impact on the puberty of male mice whose mothers were exposed to LPS (100 µg / kg) on the 12th day of pregnancy. The number of Sertoli cells were increased, whereas the body weight and the number of symplastic spermatids were decreased in offspring as compared to LPS-treated animals. Besides, IgG had a positive effect on altered hormone levels: reduced estradiol level on the 5th and 14th postnatal days and increased testosterone level on the 30th postnatal day in blood that led to an increased number of mounting attempts in sexually mature males. The cAMP-dependent pathway may be involved in the regulation of the LPS-induced inflammation. IgG reduced the increased level of cAMP in mouse peritoneal macrophages activated by LPS in vitro. IgG is able to modulate inflammation processes, but its exposure time is important.

Expression of the familial Mediterranean fever gene and activity of the C5a inhibitor in human primary fibroblast cultures

Blood ◽  
2000 ◽  
Vol 96 (2) ◽  
pp. 727-731 ◽  
Author(s):  
Yaacov Matzner ◽  
Suzan Abedat ◽  
Eli Shapiro ◽  
Shlomit Eisenberg ◽  
Ariela Bar-Gil-Shitrit ◽  
...  
Keyword(s):  
Familial Mediterranean Fever ◽  
Human Fibroblasts ◽  
Skin Fibroblasts ◽  
Sterile Inflammation ◽  
Inhibitor Activity ◽  
Primary Fibroblast ◽  
Fibroblast Cultures ◽  
M694v Mutation ◽  
Human Primary Fibroblast ◽  
Mediterranean Fever

Abstract Familial Mediterranean fever (FMF) is an inherited disease whose manifestations are acute but reversible attacks of sterile inflammation affecting synovial and serosal spaces. The FMF gene (MEFV) was recently cloned, and it codes for a protein (pyrin/marenostrin) homologous to known nuclear factors. We previously reported the deficient activity of a C5a/interleukin (IL)–8 inhibitor, a physiologic regulator of inflammatory processes, in FMF serosal and synovial fluids. We now describe the concomitant expression ofMEFV and C5a/IL-8–inhibitor activity in primary cultures of human fibroblasts. Fibroblasts grown from synovial and peritoneal tissues displayed C5a/IL-8–inhibitor activity that could be further induced with phorbol myristate acetate (PMA) and IL-1β. Very low levels of chemotactic inhibitor were evident in skin fibroblast cultures or in peritoneal and skin fibroblasts obtained from FMF patients. MEFV was expressed in peritoneal and skin fibroblasts at a lower level than in neutrophils and could be further induced by PMA and IL-1β. In the FMF cultures, the MEFV transcript carried the M694V mutation, consistent with the genetic defect found in patients with this disease. MEFV was also expressed in other cell lines that do not produce C5a/IL-8 inhibitor. These findings suggest that human primary fibroblast cultures express MEFV and produce C5a/IL-8–inhibitor activity. The interrelationship between pyrin, the MEFV product, and the C5a/IL-8 inhibitor requires further investigation.

scholarly journals Role of the 5′-untranslated region of mRNA in the synthesis of S-adenosylmethionine decarboxylase and its regulation by spermine

1994 ◽  
Vol 302 (3) ◽  
pp. 765-772 ◽  
Author(s):  
L M Shantz ◽  
R Viswanath ◽  
A E Pegg
Keyword(s):  
Secondary Structure ◽  
Untranslated Region ◽  
Fold Increase ◽  
Negative Regulator ◽  
Mrna Levels ◽  
Coding Region ◽  
Polyamine Biosynthesis ◽  
Adenosylmethionine Decarboxylase ◽  
Spermine Synthase ◽  
Synthase Inhibitor

S-Adenosylmethionine decarboxylase (AdoMetDC), a rate-limiting enzyme in polyamine biosynthesis, is regulated by polyamines at the levels of both transcription and translation. Two unusual features of AdoMetDC mRNA are a long (320 nt) 5′-untranslated region (5′UTR), which is thought to contain extensive secondary structure, and a short (15 nt) open reading frame (ORF) within the 5′UTR. We have studied the effects of altering these elements on both the expression of AdoMetDC and its regulation by n-butyl-1,3-diaminopropane (BDAP), a spermine synthase inhibitor. Human AdoMetDC cDNAs containing alterations in the 5′UTR, as well as chimaeric constructs in which the AdoMetDC 5′UTR was inserted ahead of the luciferase-coding region, were transfected into COS-7 cells. Construct pSAM320, which contains all of the 5′UTR, the AdoMetDC protein-coding region and the 3′UTR, was expressed poorly (2-fold over the endogenous activity). Deletion of virtually the entire 5′UTR, leaving nt -12 to -1, increased expression 59-fold, suggesting that 5′UTR acts as a negative regulator. The same effect was seen when the 27 nt at the extreme 5′ end were removed (pSAM293, 47-fold increase), or when the internal ORF which is present in this region was destroyed by changing the ATG to CGA (pSAM320-ATG, 38-fold increase). The expression and regulation of pSAM44 (made by deleting nt -288 to -12), which has very little predicted secondary strucutre, was very similar to that of pSAM320 indicating that the terminal 27 nt including the internal ORF rather than extensive secondary structure may be responsible for the low basal levels of AdoMetDC expression. These results, confirmed using luciferase constructs, suggest that the negative effect on expression is predominantly due to the internal ORF. Depletion of spermine by BDAP increased the expression from pSAM320 more than 5-fold without affecting AdoMetDC mRNA levels. Expression from pSAM293 was unchanged by spermine depletion, whereas that from pSAM320-ATG was increased 2.5-fold. These results indicate the presence of a spermine response element in the first 27 nt of the 5′UTR that may include but is not entirely due to the internal ORF.

Cytokine-mediated PGE2expression in human colonic fibroblasts

1998 ◽  
Vol 275 (4) ◽  
pp. C988-C994 ◽  
Author(s):  
Edward C. Kim ◽  
Yingting Zhu ◽  
Valerie Andersen ◽  
Daniela Sciaky ◽  
H. James Cao ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Cell Lines ◽  
Shape Change ◽  
Cell Types ◽  
Mrna Levels ◽  
Primary Fibroblast ◽  
Fibroblast Cultures ◽  
Epithelial Cell Lines ◽  
Factor Α

We investigated prostanoid biogenesis in human colonic fibroblasts (CCD-18Co and 5 primary fibroblast cultures) and epithelial cell lines (NCM460, T84, HT-29, and LS 174T) and the effect of PGE2 on fibroblast morphology. Cytokine-stimulated PGE2production was measured. PGH synthase-1 and -2 (PGHS-1 and -2) protein and mRNA expression were evaluated. Basal PGE2 levels were low in all cell types (0.15–6.47 ng/mg protein). Treatment for 24 h with interleukin-1β (IL-1β; 10 ng/ml) or tumor necrosis factor-α (50 ng/ml), respectively, elicited maximal 25- and 6-fold inductions of PGE2 synthesis in CCD-18Co cultures and similar results in primary fibroblast cultures; maximal inductions with IL-1β in colonic epithelial cell lines were from zero to fivefold. Treatment of CCD-18Co fibroblasts with IL-1β caused maximal 21- and 53-fold increases, respectively, in PGHS-2 protein and mRNA levels without altering PGHS-1 expression. PGE2 (0.1 μmol/l) elicited a dramatic shape change in selected fibroblasts. Colonic fibroblasts are potentially important as cytokine targets and a source of and target for colonic prostanoids in vivo.

Protective effect of artificially induced ‘hibernation’ against lethal doses of whole body x-irradiation in CF1 male mice

1959 ◽  
Vol 196 (6) ◽  
pp. 1211-1213 ◽  
Author(s):  
Sondra M. Kuskin ◽  
S. C. Wang ◽  
Roberts Rugh
Keyword(s):  
Protective Action ◽  
Lethal Dose ◽  
Lethal Effect ◽  
The Body ◽  
Whole Body ◽  
Protective Mechanism ◽  
Slight Reduction ◽  
Male Mice ◽  
Degree Of Protection ◽  
X Irradiation

Hypothermia induced by the use of neuroplegic drugs such as Hydergine, chlorpromazine or promethazine, followed by refrigeration, does not significantly enhance the protective action afforded by refrigeration alone against the lethal dose of whole body x-irradiation in CF1 male mice. The neuroplegic drugs, without refrigeration, provide a slight degree of protection, probably due to the slight reduction in the body temperature. It appears that the action of hypothermia as a protective mechanism depends not on depression of metabolism alone, but on a general depression of bodily processes. Urethane, in conjunction with refrigeration, appears to augment the lethal effect of x-irradiation in the CF1 strain of male mice.

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