We cultured the rabbit inner medullary collecting duct (IMCD) cells as monolayers on collagen-coated membrane filters, and investigated distribution of the P2Y receptors by analyzing nucleotide-induced short circuit current (Isc) responses. Exposure to different nucleotides of either the apical or basolateral surface of cell monolayers stimulated Isc. Dose-response relationship and cross-desensitization studies suggested that at least 3 distinct P2Y receptors are expressed asymmetrically on the apical and basolateral membranes. A P2Y2-like receptor, which responds to UTP and ATP, is expressed on both the apical and basolateral membranes. In addition, a uracil nucleotide receptor, which responds to UDP and UTP, but not ATP, is expressed predominantly on the apical membrane. In contrast, a P2Y1-like receptor, which responds to ADP and 2-methylthio-ATP, is expressed predominantly on the basolateral membrane. These nucleotides stimulated intracellular cAMP production with an asymmetrical profile, which was comparable to that in the stimulation of Isc. Our results suggest that the adenine and uracil nucleotides can interact with different P2Y nucleotide receptors that are expressed asymmetrically on the apical and basolateral membranes of the rabbit IMCD cells, and that both cAMP- and Ca2+-dependent signaling mechanisms underlie the stimulation of Isc.
Adenine ; Adenosine Diphosphate ; Adenosine Triphosphate ; Membranes ; Nucleotides ; Uracil ; Uracil Nucleotides ; Uridine Diphosphate ; Uridine Triphosphate

Chinese hamster ovary (CHO) cells are the preferred host cells for the production of complex recombinant therapeutic proteins. Adenine phosphoribosyltransferase (APRT) is a key enzyme in the purine biosynthesis step that catalyzes the condensation of adenine with phosphoribosylate to form adenosine phosphate AMP. In this study, the gene editing technique was used to knock out the aprt gene in CHO cells. Subsequently, the biological properties of APRT-KO CHO cell lines were investigated. A control vector expressed an enhanced green fluorescent protein (EGFP) and an attenuation vector (containing an aprt-attenuated expression cassette and EGFP) were constructed and transfected into APRT-deficient and wild-type CHO cells, respectively. The stable transfected cell pools were subcultured for 60 generations and the mean fluorescence intensity of EGFP in the recombinant CHO cells was detected by flow cytometry to analyze the EGFP expression stability. PCR amplification and sequencing showed that the aprt gene in CHO cell was successfully knocked out. The obtained APRT-deficient CHO cell line had no significant difference from the wild-type CHO cells in terms of cell morphology, growth, proliferation, and doubling time. The transient expression results indicated that compared with the wild-type CHO cells, the expression of EGFP in the APRT-deficient CHO cells transfected with the control vector and the attenuation vector increased by 42%±6% and 56%±9%, respectively. Especially, the EGFP expression levels in APRT-deficient cells transfected with the attenuation vector were significantly higher than those in wild-type CHO cells (P < 0.05). The findings suggest that the APRT-deficient CHO cell line can significantly improve the long-term expression stability of recombinant proteins. This may provide an effective cell engineering strategy for establishing an efficient and stable CHO cell expression system.
Adenine/metabolism* ; Adenine Nucleotides ; Adenine Phosphoribosyltransferase/genetics* ; Adenosine Monophosphate ; Animals ; CHO Cells ; Cricetinae ; Cricetulus ; Recombinant Proteins/genetics*

BACKGROUND: Adenosine 5'-tetraphosphate(ATPP), an endogenous nucleotide, is stored in cells and released into the extracellular space upon stimulation. Some of the biological responses to ATPP were reported, but characteristics of its receptor were not well known. Present study was conducted to investigate the effects of ATPP on mechanical contractility, resting membrane potential and action potential of rat left atrium. METHODS: Left atrium was isolated from Sprague-Dawley rat. Mechanical contraction induced by electrical field stimulation(EFS) was recorded on polygraph using force transducer. With glass microelectrodes(10 MOmega), potential difference across the membrane was measured and recorded on an oscilloscope and a polygraph. RESULTS: ATPP reduced the left atrial contractility with concentration-dependent manner. ATPP also hyperpolarized the resting membrane potential and decreased the action potential duration of the left atrial cell. Nucleotides other than ATPP, such as ATP, ADP, AMP and adenosine, have the same effect as ATPP. However, there is no difference among the nucleotides. Prior treatment of DPCPX, a P1-purinoceptor blocker, inhibited the ATPP-induced negative inotropism and changes of the membrane potential. But suramin, a nonselective P2-purinoceptor blocker, did not alter the effects of ATPP. alpha, beta methylene ADP and adenosine deaminase, which attenuates hydrolysis of adenine nucleotides and inactivates adenosine respectively, did not influence the effects of adenine nucleotides except for adenosine. CONCLUSION: ATPP reduced the mechanical contractility, hyperpolarized the resting membrance potential and decreased duration of action potential of rat left atrium. These effects were induced by ATPP directly, not by adenosine from hydrolyzed ATPP.
Action Potentials ; Adenine Nucleotides ; Adenosine Deaminase ; Adenosine Diphosphate ; Adenosine Triphosphate ; Adenosine* ; Animals ; Extracellular Space ; Glass ; Heart Atria ; Hydrolysis ; Membrane Potentials ; Membranes ; Muscle Contraction ; Nucleotides ; Rats ; Rats, Sprague-Dawley ; Receptors, Purinergic ; Suramin ; Transducers

Objective To determine the purine adenylate [adenosine triphosphate （ATP）, adenosine diphosphate （ADP）, adenosine monophosphate （AMP）] content in the muscles of both hind limbs of rats at different postmortem interval （PMI）, calculate the changes in the total adenine nucleotide （TAN） content and the adenylic-acid energy charge （AEC）, and explore their relationship with PMI. Methods Healthy rats were sacrificed by cervical dislocation and kept at 20 ℃. The muscles of their hind limbs were extracted at 0, 24, 48, 72, 96, 120, 144, and 168 h after death. Reversed-phase high performance liquid chromatography was used to determine the content of purine adenylates, the TAN and AEC of the muscles of the both hind limbs were calculated, and the related regression equations of their relationship with PMI were established. Results Within 168 h of death of rats, the trend of ATP change was different from ADP, and the content of AMP continuously increased. The TAN value gradually increased with the extension of PMI, and the AEC showed a downward trend within 168 h after death. Among them, the patterns of AEC changes with PMI were obvious, the correlation coefficient was high （ R²=0.903）, and the curve fitting relationship was good; the fitting relationship between ATP, ADP, AMP, TAN and PMI was poor （ R²=0.198-0.754）. Conclusion The postmortem change patterns of AEC provide new research ideas for PMI estimation in the forensic field.
Adenine Nucleotides ; Adenosine Monophosphate ; Animals ; Forensic Pathology ; Muscles ; Rats ; Rats, Sprague-Dawley ; Time Factors

OBJECTIVES: This study was aimed to examine whether participants of a Korean candlelight rally had correct medical information about human mad cow disease and rational attitudes about imported U.S. beef in relation to human mad cow disease. METHODS: A total of 393 face-to-face interviews were conducted, and subjects completed questions about prevalence of senile dementia and human mad cow disease in U.S. and whether they will eat U.S. beef even if no cases of human mad cow disease occurred in the U.S. or if the chance of being affected with human mad cow disease was lower than dying in a plane crash. RESULTS: Correct answer rates to the questions about prevalence of senile dementia and human mad cow disease were 28.2% and 36.1%, respectively. A majority of respondents answered that they would not eat U.S. beef even if there were no reported cases of human mad cow disease in the U.S. or if their chance of being affected with human mad cow disease was lower than dying in a plane crash (75.6% and 86.0%, respectively). CONCLUSION: At least 64.4% of participants had incorrect medical information about human mad cow disease, and their attitudes about imported U.S. beef may be rooted in emotion rather than fact.
Adenine Nucleotides ; Alzheimer Disease ; Animals ; Cattle ; Surveys and Questionnaires ; Encephalopathy, Bovine Spongiform ; Humans ; Korea ; Mycophenolic Acid ; Prevalence

PURPOSE: The authors sought to assess the usefulness of navigation as opposed to the conventional method by analyzing the radiographic results obtained from subjects who underwent total knee arthroplasty for knees that were accompanied with anatomic variations. MATERIALS AND METHODS: The study subjects were selected from 53 patients (a total 72 cases: 43 were treated by the conventional method and 29 were treated by the navigational method) who exhibited radiographic evidence of distal femoral varus (<0degrees) or proximal tibial varus (>2degrees). The coronal femoral component angle (alpha) and the coronal tibial component angle (beta) were measured, and the femoral component position in relation to the mechanical axis (theta) and the post-operative weight-bearing mechanical axis difference (MAD) were compared and analyzed. RESULTS: The navigation method showed significant better results in terms of the alpha, theta and MAD (p<0.05). Among the outliers greater than 3degrees, a statistically significant difference was shown only for the MAD (p=0.030). CONCLUSION: Navigation surgery is useful in terms of the femoral component's position in the coronal plane and limb alignment in the osteoarthritic knee that is accompanied by distal femoral varus or proximal tibial varus.
Adenine Nucleotides ; Anatomic Variation ; Arthroplasty ; Axis, Cervical Vertebra ; Extremities ; Humans ; Knee ; Mycophenolic Acid ; Weight-Bearing

OBJECTIVE: To investigate differences of the postural control in unstable sitting position between elderly and young adults. METHOD: Twenty five healthy elderly and twenty five healthy young adults were included. The evaluation system for postural control consisted of unstable plate, frame, safety harness, monitor and computer. Subjects sat on an unstable plate with arms crossed. Using two tilt sensor and postural control software in unstable platform measured the center of pressure (COP) of subject. COP sway (COP was maintained on the center circle and the distance from the central location for 30 sec) time and mean absolute deviation (MAD), COP maintaining (COP was maintained on the desired target in anterior, posterior, left or right directions during 30 sec) time and MAD, COP moving time (the time required to move the COP to desired target location away from center), COP sine curve maintaining (COP was maintained on the circle on moving sine curve during 30 sec) time and MAD were recorded in both groups. Each subject performed three trials and the mean value of the trials was used for analysis. RESULTS: In static evaluation, there was no significant difference in COP sway between two groups. In dynamic evaluations, elderly showed significantly decreased maintaining time in all four directions, decreased sine curve trace and increased moving time in all eight directions (p<0.001). CONCLUSION: Elderly revealed significantly impaired dynamic sitting postural control, regardless of directions. It might be related to decreased movement and proprioception of trunk.
Adenine Nucleotides ; Aged ; Arm ; Humans ; Mycophenolic Acid ; Organothiophosphorus Compounds ; Proprioception ; Young Adult

PURPOSE: The purpose of this study was to evaluate clinical results and accuracy of femoral cutting in the coronal plane in total knee arthroplasty (TKA) using a fixed length intramedullary guide. MATERIALS AND METHODS: From 2005 to 2008, 101 patients (154 knees) underwent TKA (NexGen LPS implant). The minimal follow-up period was 3 years (mean, 4.4 years). The patients were divided into two groups (group 1, 94< or =alpha angle<98; group 2, 94>alpha, 98< or =alpha). Mechanical axis deviation (MAD), patellar tilting angle (PTA), Knee Society Knee Score (KSKS), and Knee Society Function Score (KSFS) were measured in both groups. RESULTS: There were 120 knees in group 1 and 34 knees in group 2. There was no significant intergroup difference in the postoperative MAD (group 1, 1.59degrees; group 2, 1.91degrees). The number of outliers with > or =2degrees MAD was 65 in group 1 and 24 in group 2. The mean PTA, KSKS, and KSFS were 10.17degrees, 96.0, and 96.6, respectively, in group 1 and 11.58degrees, 84.5, and 85.5, respectively, in group 2. CONCLUSIONS: The percentage of coronal alignment outliers was relatively high (34 in 154 cases, 22%) after TKA using a fixed length intramedullary guide. However, there was no statistically significant intergroup difference in clinical results (KSKS, p=0.67; KSFS, p=0.56).
Adenine Nucleotides ; Arthroplasty ; Axis, Cervical Vertebra ; Follow-Up Studies ; Humans ; Knee ; Mycophenolic Acid

Glucose and lipid metabolism is the most fundamental metabolic activity of higher organisms. This process is affected by both genetic polymorphisms and environmental factors. Excessive uptake and accumulation of lipids lead to obesity and disorder of glucose metabolic homeostasis characterized by insulin resistance and hyperglycemia, suggesting that the cross-regulation between lipid and glucose metabolism happens precisely at organ, cellular and molecular levels by known mechanisms. Adenine nucleotides and their metabolites have emerged as mediators in the mutual regulation of glucose and lipid metabolism. This review summarizes the roles of purinergic signaling induced by fatty acids in glucose metabolism and the development of type 2 diabetes.
Adenine Nucleotides ; Diabetes Mellitus, Type 2 ; Glucose ; Homeostasis ; Humans ; Insulin Resistance ; Lipid Metabolism

OBJECTIVES: The purposes of this study were to estimate the effect of mandibular advancement device (MAD) and to evaluate the influence of the advancement amount of mandible in the application of MAD for obstructive sleep apnea (OSA) patients. METHODS: From the patients who were diagnosed as OSA by polysomnographic study at Seoul National University Bundang Hospital from January 2007 to February 2009, the patients who chose MAD as treatment option were included in this study. All the patients' data including clinical records and polysomnographic studies (both pre- and post-treatment) were reviewed and analyzed. RESULTS: Successful results were obtained in 65 patients of 86 patients (75.6%). In the follow-up period, mild discomfort of anterior teeth or temporomandibular joint (TMJ) were described in 28 patients, especially in the cases the amount of mandibular advancement were more than 7.0 mm. There was no direct relationship between the amount of mandibular advancement and clinical outcome. CONCLUSION: MAD was effective treatment option for the OSA patients regardless of severity. For the prevention of potential dental complications, the amount of mandibular advancement should be considered at the time of MAD treatment.
Adenine Nucleotides ; Follow-Up Studies ; Humans ; Mandible ; Mandibular Advancement ; Mycophenolic Acid ; Sleep Apnea, Obstructive ; Temporomandibular Joint ; Tooth