BACKGROUND: The endothelium-dependent vasorelaxation has been largely accounted for by the release of nitric oxide (NO). Three distinct isoforms of NO synthases (NOS) have been characterized, i.e., brain(bNOS), inducible (iNOS), and endothelial constitutive (ecNOS). Although hypertension hasbeen associated with a vascular endothelial dysfunction, changes in the vascular expression of NOS isoforms have not been established. The present study was aimed at exploring the vascular expression of NOS isozymes in hypertension. MATERIAL AND METHOD: Two-kidney, one clip (2K1C) and deoxycorticosterone acetate (DOCA)-salt hypertension were induced in rats. The expression of different NOS isozymes in the thoracic aorta was determined by Western blot analysis. The vascular tissue contents of nitrites were measured by colorimetric assay. RESULT: Arterial blood pressure was significantly higher in experimental groups of 2K1C and DOCA-salt rats compared with their corresponding control rats. The vascular expression of bNOS as well as that of ecNOS was decreased in both models of hypertension. iNOS was not changed in DOCA-salt hypertension, but was also decreased in 2K1C hypertension. The vascular contents of nitrites were significantly decreased in DOCA-salt as well as in 2K1C hypertension. CONCLUSION: These results suggest that 2K1C and DOCA-salt hypertension are associated with decreases in the vascular expression of NOS isozymes and nitrite contents.
Animals ; Aorta, Thoracic ; Arterial Pressure ; Blotting, Western ; Desoxycorticosterone ; Hypertension* ; Isoenzymes ; Nitric Oxide Synthase* ; Nitric Oxide* ; Nitrites ; Protein Isoforms ; Rats ; Vasodilation

No abstract available.
Gingiva* ; Periodontitis ; Protein Isoforms* ; Superoxide Dismutase* ; Superoxides*

Clusterin (CLU) is a multifunctional glycoprotein that has secretory and nuclear isoforms. The two isoforms are known to play opposite roles in cell survival/death. In this review, we summarize recent progress on the pro-apoptotic function of nuclear CLU in vitro and in vivo and discuss previous reports on the role of CLU in brain damage and neurodegeneration.
Apoptosis ; Brain ; Clusterin ; Glycoproteins ; Protein Isoforms

Acute promyelocytic leukemia (APL) is characterized by a specific t (15;17) translocation which produce a PML/RAR-alpha fusion messenger RNA and by effectiveness of all-trans retinoic acid (ATRA) differentiation therapy. Breakpoints within PML intron 3 (bcr 3) produce a short PML/RAR-alpha isoform (S-isoform), whereas breakpoints within PML intron 6 (bcr 1) result in a longer form (L-isoform). Additionally, breakpoints within PML exon 6 (bcr 2) make a variable length transcript (V-isoform) in a small number of patients. The influence of breakpoint site on patient outcome remains controversial. Previous reports showed that patients with S-isoform have an increased incidence of clinical relapse and shorter survival compared to those with L-isoform. Others reported no difference in DFS between these patients groups. In this issue, Lee et al. reported that there were 58 L-isoform (62.1%), 32 S-isoform (34.0%), 4 V-isoform (4.3%) and, no significant prognostic factor for EFS from induction therapy using anthracycline plus ATRA among 94 patients with APL. They concluded pretreatment clinical characteristics and treatment outcomes were not significantly different according to PML/RAR-alpha isoform types in this induction group. Recently, it was reported that FLT3/ITD mutation was frequently associated with S-isoform and with the M3v form of leukemia and CNS relapse in APL was mostly related to S-isoform. With previous studies including this article, outcomes of different types of PML/RAR-alpha isoforms are not conclusive. Future researches need to be focused not only on clinical outcomes of different types of PML/RAR-alpha isoforms, but also clinical relevance of PML/RARA-alpha mRNA isoforms with other prognostic factors and particular clinical characteristics.
Exons ; Humans ; Incidence ; Introns ; Leukemia ; Leukemia, Promyelocytic, Acute ; Protein Isoforms ; Recurrence ; RNA Isoforms ; RNA, Messenger ; Tretinoin

OBJECTIVES: Osteoclasts are multinucleated giant cells which can resorb bone and differentiated from hematopoietic cells. We have previously reported murine osteoclast-associated receptor (OSCAR) may be an important bone-specific regulator of osteoclast differentiation. We have cloned soluble form of human OSCAR (hOSCAR) and examined the role of hOSCAR on osteoclast differentiation. METHODS: Osteoclast differentiation was induced by treatment with macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor kappa B ligand (RANKL) and tartrate-resistant acid phosphatase (TRAP) staining and pit formation were performed. Expression was measured by flow cytometry analysis, Northern and Western blot analysis. RESULTS: hOSCAR is expressed in osteoclast cells and involved in the differentiation of osteoclasts from peripheral blood mononuclear cells (PBMC). Two alternatively spliced forms (soluble hOSCAR [hOSCAR-S]) of hOSCAR were identified from osteoclasts complementary deoxyribonucleic acid (cDNA) library derived from PBMC. Putative transmembrane domain was not found in hOSCAR-S forms and it suggested that these forms might be secreted from osteoclast cells. These secreted forms of hOSCAR attenuated RANKL-induced osteoclast formation and bone resorption. CONCLUSIONS: Human osteoclasts express at least five different OSCAR messenger ribonucleic acid (mRNA) isoforms which could play different regulatory roles for differentiation. The secreted forms of hOSCAR might be a negative regulator of membrane-bounded forms of OSCAR.
Acid Phosphatase ; Alternative Splicing ; Blotting, Western ; Clone Cells ; DNA ; Flow Cytometry ; Giant Cells ; Humans ; Isoenzymes ; Macrophage Colony-Stimulating Factor ; Osteoclasts ; Protein Isoforms ; RANK Ligand ; RNA

OBJECTIVES: Osteoclasts are multinucleated giant cells which can resorb bone and differentiated from hematopoietic cells. We have previously reported murine osteoclast-associated receptor (OSCAR) may be an important bone-specific regulator of osteoclast differentiation. We have cloned soluble form of human OSCAR (hOSCAR) and examined the role of hOSCAR on osteoclast differentiation. METHODS: Osteoclast differentiation was induced by treatment with macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor kappa B ligand (RANKL) and tartrate-resistant acid phosphatase (TRAP) staining and pit formation were performed. Expression was measured by flow cytometry analysis, Northern and Western blot analysis. RESULTS: hOSCAR is expressed in osteoclast cells and involved in the differentiation of osteoclasts from peripheral blood mononuclear cells (PBMC). Two alternatively spliced forms (soluble hOSCAR [hOSCAR-S]) of hOSCAR were identified from osteoclasts complementary deoxyribonucleic acid (cDNA) library derived from PBMC. Putative transmembrane domain was not found in hOSCAR-S forms and it suggested that these forms might be secreted from osteoclast cells. These secreted forms of hOSCAR attenuated RANKL-induced osteoclast formation and bone resorption. CONCLUSIONS: Human osteoclasts express at least five different OSCAR messenger ribonucleic acid (mRNA) isoforms which could play different regulatory roles for differentiation. The secreted forms of hOSCAR might be a negative regulator of membrane-bounded forms of OSCAR.
Acid Phosphatase ; Alternative Splicing ; Blotting, Western ; Clone Cells ; DNA ; Flow Cytometry ; Giant Cells ; Humans ; Isoenzymes ; Macrophage Colony-Stimulating Factor ; Osteoclasts ; Protein Isoforms ; RANK Ligand ; RNA

We found that non-small-cell lung cancer (NSCLC) cells express high levels of multiple aldehyde dehydrogenase (ALDH) isoforms via an informatics analysis of metabolic enzymes in NSCLC and immunohistochemical staining of NSCLC clinical tumor samples. Using a multiple reaction-monitoring mass spectrometry analysis, we found that multiple ALDH isozymes were generally abundant in NSCLC cells compared with their levels in normal IMR-90 human lung cells. As a result of the catalytic reaction mediated by ALDH, NADH is produced as a by-product from the conversion of aldehyde to carboxylic acid. We hypothesized that the NADH produced by ALDH may be a reliable energy source for ATP production in NSCLC. This study revealed that NADH production by ALDH contributes significantly to ATP production in NSCLC. Furthermore, gossypol, a pan-ALDH inhibitor, markedly reduced the level of ATP. Gossypol combined with phenformin synergistically reduced the ATP levels, which efficiently induced cell death following cell cycle arrest.
Adenosine Triphosphate ; Aldehyde Dehydrogenase* ; Cell Cycle Checkpoints ; Cell Death ; Energy Metabolism* ; Gossypol ; Humans ; Informatics ; Isoenzymes ; Lung ; Lung Neoplasms ; Mass Spectrometry ; NAD ; Phenformin ; Protein Isoforms

The endolymph and perilymph of the inner ear have unique ionic composition and electrical potential. It is widely accepted that normal auditory function depends on them and Na/K-ATPase plays a central role in production and maintenance of them. The distribution of five Na/K-ATPase subunit isoform (alpha1, alpha2, alpha3, beta1, and beta2) in rat inner ear was determined by immunohistochemistry after decalcifying the temporal bone with Gooding and Stewart's solution. In the cochlear regions, Na/K-ATPase alpha1beta1 isozyme was abundantly expressed in the infrastrial fibrocytes, suprastrial fibrocytes, spiral prominence, outer sulcus cells and spiral ganglion, and also detected in cochlear nerve and interdental cells. alpha1beta2 isozyme was abundantly expressed in all layers of stria vascularis and alpha3beta1 isozyme was detected in cochlear nerve and spiral ganglion. alpha3beta2 isozyme was expressed in spiral ganglion. In vestibular regions, Na/K-ATPase alpha1b1 isozyme was expressed in macular sacculi hair cell, transitional cells of ampulla, and vestibular ganglion, and alpha1b2 isozyme was abundantly expressed in ampullary dark cells and transitional cells and vestibular ganglion. a3b1 isozyme was abundantly expressed in crista ampularis, macula utriculi, and macula sacculi hair cells, and also moderately detected in ampullary, utricular, and saccular nerves, and vestibular ganglion. alpha3beta2 isozyme also detected in ampullary, utricular, and saccular nerves, and vestibular ganglion. But, alpha2beta1 and alpha2beta2 isozymes were not detected in any regions of inner ear. These findings suggest the possibility of four unique Na/K-ATPase isozymes deferentially expressed among the various cell types of the inner ear. This structural diversity imparts considerable biological versatility to the Na/K-ATPase and would be provided the explanations for the differences in fluid and ion transport and its regulation among the inner ear regions.
Animals ; Cochlear Nerve ; Ear, Inner* ; Endolymph ; Ganglion Cysts ; Hair ; Immunohistochemistry ; Ion Transport ; Isoenzymes ; Perilymph ; Protein Isoforms* ; Rats* ; Spiral Ganglion ; Stria Vascularis ; Temporal Bone

PURPOSE: To investigate the cellular protective effects of hypoxic preconditioning against oxidative stress in a staurosporine-differentiated RGC-5 cell line and the relevance of protein kinase C subtype expression. METHODS: The minimum staurosporine concentration and exposure time necessary to morphologically fully differentiate RGC-5 cells were determined. Cytotoxic injury was provided by oxidative stress with 800 micrometer hydrogen peroxide (H2O2) for 15 hours to morphologically fully-differentiated cells. The cytoprotective effect of hypoxic preconditioning was found by exposing the cell line to 0.3% oxygen for different periods of time. Quantifiable changes in the expression of mRNAs and proteins of the isoenzymes alpha, beta, gamma, delta, epsilon, zeta of protein kinase C were determined before and after 1, 2, 15, and 24 hours of hypoxic preconditioning. RESULTS: Axonal growth in RGC-5 cells after the induction of differentiation with staurosporine caused these cells to resemble neurons. The minimal concentration and exposure time to staurosporine that evoked full differentiation of RGC-5 cells was exposure to 2 micrometer staurosporine for 1 hour. An LDH assay demonstrated that hypoxic preconditioning had neuroprotective effects against hydrogen peroxide-induced oxidative stress. Protein and mRNA levels of PKC isoforms alpha and epsilon increased after preconditioning. CONCLUSIONS: Hypoxic preconditioning of staurosporine-differentiated RGC-5 cells had a cytoprotective effect against oxidative stress. The associated increase of mRNA and proteins of PKC isoenzymes alpha and epsilon suggest some functional relevance of these isoenzymes to the cytoprotective effects conferred by hypoxic preconditioning.
Axons ; Cell Line ; Hydrogen ; Hydrogen Peroxide ; Ischemic Preconditioning ; Isoenzymes ; Neurons ; Neuroprotective Agents ; Oxidative Stress ; Oxygen ; Protein Isoforms ; Protein Kinase C ; Protein Kinases ; Proteins ; Retinal Ganglion Cells ; Retinaldehyde ; RNA, Messenger ; Staurosporine

Ischemic preconditioning (IPC) is a phenomenon that a brief episode of ischemia to a tissue renders the tissue resistance from a subsequent prolonged ischemia. It is generally accepted that this protection is a receptor-mediated process, and is realized via signal transduction pathways. Protein kinase C (PKC), known to play key regulatory roles in cellular processes, has been proposed as a primary cellular mediator of preconditioning. However, the role of PKC in eliciting cardioprotection remains controversial. The evidences for the 'PKC hypothesis' of preconditioning in various tissue and organs are summarized. Especially in intestine, a brief ischemia induced a reversible epithelial injury to the jejunum that is associated with activation of several PKC isoforms. Injury induced by an additional period of ischemia is reduced by the prior IPC, and this effect is abolished by non-selective PKC inhibition but not by a selective inhibitor of cPKC/or PKCdelta. This result suggest that activation of nPKC isoform (especially PKCepsilon) during and following ischemic insults may play an important role in protection against I/R injury in the intestine, and this mechanism is identical with previous study in heart tissue.
Heart ; Intestines ; Ischemia ; Ischemic Preconditioning* ; Jejunum ; Protein Isoforms ; Protein Kinase C* ; Protein Kinases* ; Signal Transduction