• Journal of Veterinary Science
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• v.24 no.2
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• pp.26.1-26.11
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• 2023

Background: Angiotensin-converting enzyme inhibitor (ACEi) inhibits the catalysis of angiotensin I to angiotensin II and the degradation of substance P (SP) and bradykinin (BK). While the possible relationship between ACEi and SP in nociceptive mice was recently suggested, the effect of ACEi on signal transduction in astrocytes remains unclear. Objectives: This study examined whether ACE inhibition with captopril or enalapril modulates the levels of SP and BK in primary cultured astrocytes and whether this change modulates PKC isoforms (PKCα, PKCβI, and PKCε) expression in cultured astrocytes. Methods: Immunocytochemistry and Western blot analysis were performed to examine the changes in the levels of SP and BK and the expression of the PKC isoforms in primary cultured astrocytes, respectively. Results: The treatment of captopril or enalapril increased the immunoreactivity of SP and BK significantly in glial fibrillary acidic protein-positive cultured astrocytes. These increases were suppressed by a pretreatment with an angiotensin-converting enzyme. In addition, treatment with captopril increased the expression of the PKCβI isoform in cultured astrocytes, while there were no changes in the expression of the PKCα and PKCε isoforms after the captopril treatment. The captopril-induced increased expression of the PKCβI isoform was inhibited by a pretreatment with the neurokinin-1 receptor antagonist, L-733,060, the BK B1 receptor antagonist, R 715, or the BK B₂ receptor antagonist, HOE 140. Conclusions: These results suggest that ACE inhibition with captopril or enalapril increases the levels of SP and BK in cultured astrocytes and that the activation of SP and BK receptors mediates the captopril-induced increase in the expression of the PKCβI isoform.

• Archives of Pharmacal Research
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• v.21 no.6
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• pp.707-711
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• 1998

The (Na,K)ATPase is responsible for generating the ionic gradients and membrane potentials by the exchange of intracellular $Na^+$ for $K^+$. It has been recentl y shown that (Na,K)ATPase is involved in the exocytic pathway of basic fibroblast growth factor (bFGF), although it is not known that bFGF is secreted to the outside of cell through direct interaction with (Na,K) ATPase. To understand the role for (Na,K)ATPase in the secretary pathway of bFGF, we have sought to identify the cytoplasmic domains of the alpha1 isoform of (Na,K)ATPase interacting with bFGF by yeast two-hybrid system. We have also investigated the interaction between the alpha2 isoform of (Na,K)ATPase and bFGF to find out whether the interaction is isoform-specific. We found that none of the cytoplasmic domains of (Na,K)ATPase isoforms interacted with bFGF. The result suggests that the interaction between bFGF and (Na,K)ATPase might be indirect, thus requiring other proteins which are involved in the formation of protein complexes for the interaction, although we cannot exclude the possibility that the interaction requires the element of the whole alpha subunit structure that was not present in the isolated alpha subunit cytoplasmic domains.

• Journal of fish pathology
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• v.24 no.3
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• pp.269-278
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• 2011

Gene expression of two glyceraldehyde-3-phosphate dehydrogenase (GAPDH) paralogs was examined during Edwardsiella tarda challenge and heavy metal exposures in mud loach (Misgurnus mizolepis; Cypriniformes) kidney and spleen. Transcription of the two mud loach GAPDH paralogs (mlGAPDH-1 and mlGAPDH-2) was significantly modulated by these stimulatory challenges in an isoform-dependent manner. Based on the real-time RT-PCR analysis, the mlGAPDH-2 transcripts were more preferentially induced by E. tarda challenge, whereas the mlGAPDH-1 transcripts were proven to show more inducibility in response to heavy metal exposure using Cd, Cu, Mn and Zn at $5{\mu}M$. Their isoform-specific response patterns were closely in accordance with the TF binding profiles in promoter and intron-1 of the two mlGAPDH isoforms, in which the mlGAPDH-2 has more binding sites for immune-related transcription factors than mlGAPDH-1 while the mlGAPDH-1 possesses exclusively metal responsive elements in its intron. Collectively, the mlGAPDHs are potentially involved in cellular pathways independent of glycolysis and the two GAPDH paralogs might undergo functional diversification or subfunctionalization at least at the transcription level.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.300-300
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• 1994

$Na^{+}$,$K^{+}$-ATPase activity, $Na^{+}$-dependent phosphorylalion, and [$^3$〕ouabain binding in sarcolemma prepared from 4 week old spontaneously hypertensive rat(SHR) ventricles were compared to the same parameters in sarcolemma from age matched normotensive Wistar-Kyoto(WKY) rat ventricles to examine whether the reduced myocardial $Na^{+}$-pump activity in SHR is an inherited enzymatic defect or a second phenomenon due to sustained hypertension. The total body weights, ventricular weights, and blood pressures were the same for SHR and WKY. No significant differences in sarcolemmal protein content and protein recovery were noted between the two pups. Sarcolemma isolated from SHR ventricles showed sigificantly less $Na^{+}$,$K^{+}$-ATPase activity and number of phosphorylation sites when compared to snrcolemma 1mm the WKY ventricles. Equilibrium binding of [$^3H$〕ouabain and the turnovcr number of myocardial $Na^{+}$,$K^{+}$-ATPast however, wee the same for both groups. These results. indicate that the low affinity(${\alpha}$, or ${\alpha}_1$) isoform for ouabain is reduced in SHR compared to WKY but that the high affinity(${\alpha}$+, or ${\alpha}_2$) isoform is the same in ventricles of SHR and WKY. The reduced amount of at isoform of the $Na^{+}$,$K^{+}$-ATPasc in prehypertensive SHR ventricles may play some pole in the development of hypertension.

• BMB Reports
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• v.38 no.1
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• pp.28-33
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• 2005

A gene coding a novel isoform of carbamyl phosphate synthetase I (CPS1) was cloned from a human testicular library. As shown by cDNA microarray hybridization, this gene was expressed at a higher level in human adult testes than in fetal testes. The full length of its cDNA was 3831 bp, with a 3149 bp open reading frame, encoding a 1050-amino-acid protein. The cDNA sequence was deposited in the GenBank (AY317138). Sequence analysis showed that it was homologous to the human CPS1 gene. The putative protein contained functional domains composing the intact large subunit of carbamoyl phosphate synthetase, thus indicated it has the capability of arginine biosynthesis. A multiple tissue expression profile showed high expression of this gene in human testis, suggesting the novel alternative splicing form of CPS1 may be correlated with human spermatogenesis.

• Parasites, Hosts and Diseases
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• v.51 no.5
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• pp.497-502
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• 2013

Autophagy-related protein 8 (Atg8) is an essential component of autophagy formation and encystment of cystforming parasites, and some protozoa, such as, Acanthamoeba, Entamoeba, and Dictyostelium, have been reported to possess a type of Atg8. In this study, an isoform of Atg8 was identified and characterized in Acanthamoeba castellanii (AcAtg8b). AcAtg8b protein was found to encode 132 amino acids and to be longer than AcAtg8 protein, which encoded 117 amino acids. Real-time PCR analysis showed high expression levels of AcAtg8b and AcAtg8 during encystation. Fluorescence microscopy demonstrated that AcAtg8b is involved in the formation of the autophagosomal membrane. Chemically synthesized siRNA against AcAtg8b reduced the encystation efficiency of Acanthamoeba, confirming that AcAtg8b, like AcAtg8, is an essential component of cyst formation in Acanthamoeba. Our findings suggest that Acanthamoeba has doubled the number of Atg8 gene copies to ensure the successful encystation for survival when 1 copy is lost. These 2 types of Atg8 identified in Acanthamoeba provide important information regarding autophagy formation, encystation mechanism, and survival of primitive, cyst-forming protozoan parasites.

• Fisheries and Aquatic Sciences
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• v.14 no.4
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• pp.303-310
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• 2011

We characterized a cytoplasmic actin gene locus in greenling Hexagrammos otakii (Scorpaeniformes). Genomic clones isolated from the greenling DNA library contained two homologous cytoplasmic actin gene copies (HObact2.1 and HObact2.2) in a tail-to-head orientation. Their gene structure is characterized by six translated exons and one non-translated exon. Exon-intron organization and the nucleotide sequences of the two actin gene isoforms are very similar. However, only the HObact2.1 isoform contains microsatellite-like, dinucleotide repeats in the 5'-flanking region (named HOms2002) and intron 1 following the non-translated exon 1 (named HOms769). One microsatellite locus (HOms769) was highly polymorphic while the other (HOms2002) was not. Based on bioinformatic analysis, different transcription factor binding motifs are related to stress and immune responses in the two actin isoforms. Semiquantitative and real-time reverse transcription-PCR assays showed that both isoform transcripts were detectable ubiquitously in all the tissues examined. However, the basal expression levels of each isoform varied across tissues. Overall, the two isoforms showed a similar, but not identical, expression pattern. Our data suggest that the cytoplasmic actin genes may be the result of a recent duplication event in the greenling genome, which has not experienced significant subfunctionalization in their housekeeping roles.

• Proceedings of the Ginseng society Conference
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• 1998.06a
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• pp.244-252
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• 1998

Ginsenoside Rh, (G-Rh,) from Panax ginseng induced morphological features of apoptosis and DNA fragmentation as a biochemical marker of apoptosis confirmed by TUNEL reaction and agarose gel electrophoresis in human neuroblastoma SK-N-BE(2) and rat glioma C6Bu-1 cells During apoptosis by G-Rh2, protein kinase C (PKC) isoforms were analysed by immunoblotting. In SK-N-BE(2) cells, the levels of a, p and ${\gamma}$ subtypes were increased by undergoing apoptosis, while PKC e isoform increased early in treatment (3 h and 6 h). In addition, PKC s isoform gradually decreased during apoptosis by G-Rh2 and PKC $\theta$ isoform was detected in neither untreated- nor G-Rh1-treated SK-N-BE(2) cells (data not shown). However, no significant changes in the level of S and s isoforms were observed in C6Bu-1 cells undergoing apoptosis by G-Rh2. These results suggest that PKC subtypes may play differential roles in apoptotic signal pathways and their roles can be cell type-specific in apoptosis induced by G-Rh2.

• Parasites, Hosts and Diseases
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• v.57 no.3
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• pp.249-256
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• 2019

Steroids are commonly used in patients with eosinophilic meningitis caused by A. cantonensis infections. The mechanism steroids act on eosinophilic meningitis remains unclear. In this mouse experiments, expressions of 14-3-3 isoform ${\beta}$ and ${\gamma}$ proteins significantly increased in the CSF 2-3 weeks after the infection, but not increasedin the dexamethasone-treated group. Expression of 14-3-3 ${\beta}$, ${\gamma}$, ${\varepsilon}$, and ${\theta}$ isoforms increased in brain meninges over the 3-week period after infection and decreased due to dexamethasone treatment. In conclusion, administration of dexamethasone in mice with eosinophilic meningitis decreased expressions of 14-3-3 isoform proteins in the CSF and in brain meninges.

• Korean Journal of Food Science and Technology
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• v.35 no.5
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• pp.791-796
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• 2003

Polyphenol oxidase isoforms were purified from the lotus roots using 50% acetone precipitation, conventional chromatographies of Q-Sepharose and hydrophobic interaction, and high performance liquid chromatographies of Mono-Q and Superdex 75 gel-filtration. Molecular mass of a purified PPO isoform (LPIII-2) was determined to be 56 kDa using gel-filtration chromatography. The active form of LPIII-2 appeared to bea heterodimer, as purified LPIII-2 on SDS-PAGE gel showed two bands that were determined to be 28 kDa and 26 kDa. To further characterize PPO, partially purified PPO isoforms (LP-II, LP-III) were obtained from Q-Sepharose anion-exchange chromatography. In substrate specificity, the partially purified PPO isoform LP-II showed a high affinity to catechol, while LP-III showed a high affinity to pyrogallol. The optimum pH of LP-II and LP-III was pH 7.0. Interestingly, the partially purified PPO isoforms showed high activities at low temperatures $(0{\sim}5^{\circ}C)$, and as temperatures rose, the activities decreased. Both PPO isoforms were stable at $40^{\circ}C$ and were inactivated by incubation at $60^{\circ}C$ for 40 min.