LMBD1 protein participates in cell mitosis by regulating microtubule assembly.

LMBD1 was previously demonstrated to regulate the endocytosis of insulin receptor on the cell surface and to mediate the export of cobalamin from the lysosomes to the cytosol, but little is known about its function in mitosis. In this study, interactome analysis data indicate that LMBD1 is involved in cytoskeleton regulation. Both immunoprecipitation and GST pulldown assays demonstrated the association of LMBD1 with tubulin. Immunofluorescence staining also showed the colocalization of LMBD1 with microtubule in both interphase and mitotic cells. LMBD1 specifically accelerates microtubule assembly dynamics in vitro and antagonizes the microtubule-disruptive effect of vinblastine. In addition, LMBRD1-knockdown impairs mitotic spindle formation, inhibits tubulin polymerization, and diminishes the mitosis-associated tubulin acetylation. The reduced acetylation can be reversed by ectopic expression of LMBD1 protein. These results suggest that LMBD1 protein stabilizes microtubule intermediates. Furthermore, embryonic fibroblasts derived from Lmbrd1 heterozygous knockout mice showed abnormality in microtubule formation, mitosis, and cell growth. Taken together, LMBD1 plays a pivotal role in regulating microtubule assembly that is essential for the process of cell mitosis.

Incidence of Unintentional Flow of Contrast into the Facet Joints During Fluoroscopy-Guided Cervical Interlaminar Epidural Injections: A Retrospective Cohort Study.

OBJECTIVE: We sometimes encounter unintentional flow of contrast into the facet joints during cervical interlaminar epidural injection, which leads to false-positive epidural injection. The purposes of this study were to evaluate the rate of facet flow of contrast and to investigate various factors associated with injection into the space of Okada during fluoroscopy-guided cervical interlaminar epidural injection. SETTING AND SUBJECTS: Images from consecutive cases of fluoroscopy-guided cervical interlaminar epidural injection performed at a single institution between July 2015 and July 2018 were obtained and reviewed. METHODS: Cases of epidural injection were classified as either facet flow or no facet flow. Multivariate logistic regression was used to identify the predictive factors of unintended injection into the Okada space. RESULTS: A total of 2,006 cases were included. Intra-articular flow was identified in 6.0% of cases (121/2,006). All cases of flow of contrast into the facet joints were recognized, and appropriate epidurograms were obtained during the procedures. The highest rate of unintended facet flow of the contrast (10.1%, 44/436) occurred at C5-6. Cervical interlaminar epidural injection at C5-6 and above (adjusted odds ratio [aOR] = 1.929, P = 0.001) and the paramidline approach for epidural injection (aOR = 2.427, P < 0.001) were associated with injection into the space of Okada. CONCLUSIONS: We detected injection into the space of Okada during fluoroscopy-guided cervical interlaminar epidural injection in 6.0% of procedures. Cervical interlaminar epidural injection at C5-6 and above and the paramidline approach for epidural injection were positive predictors of unintentional facet flow of the contrast.

Quantitative display of the redox status of proteins with maleimide-polyethylene glycol tagging.

Cysteine oxidation, either biologically reversible or irreversible, is the main posttranslational modification associated with redox signaling and oxidative stress. Maleimide-polyethylene glycol (m-PEG) has been used to detect reversibly oxidized proteins by reacting to the reduced cysteine residues leading to mobility shift in immunoblots; a method called PEG-switch. With PEG-switch, both reduced and oxidized proteins can be observed on the same immunoblot simultaneously, providing a simple quantitative measurement for protein thiol modifications. In this report, we optimized the assay conditions and exploited the applications of PEG-switch in quantitation of the extent of protein thiol oxidation in cells in response to H2 O2 and insulin. In addition, we have proposed a redox scoring system for measuring the redox status of any given protein from the m-PEG immunoblot. Our results provided quantitative data showing that two cysteine residues of protein tyrosine phosphatase 1B are prone to oxidation following insulin treatment in cultured HeLa cells.

Mass spectrometric imaging of metabolites in kidney tissues from rats treated with furosemide.

In the kidney, metabolic processes are different among the cortex (COR), outer medulla (OM), and inner medulla (IM). Using matrix-assisted laser desorption/ionization (MALDI) and imaging mass spectrometry (IMS), we examined the change of metabolites in the COR, OM, and IM of the rat kidney after furosemide treatment compared with vehicle-treated controls. Osmotic minipumps were implanted in male Sprague-Dawley rats to deliver 12 mg·day(-1)·rat(-1) of furosemide. Vehicle-treated (n = 14) and furosemide-treated (furosemide rats, n = 15) rats in metabolic cages received a fixed amount of rat chow (15 g·220 g body wt(-1)·day(-1) for each rat) with free access to water intake for 6 days. At day 6, higher urine output (32 ± 4 vs. 9 ± 1 ml/day) and lower urine osmolality (546 ± 44 vs. 1,677 ± 104 mosmol/kgH2O) were observed in furosemide rats. Extracts of COR, OM, and IM were analyzed by ultraperformance liquid chromatography coupled with quadrupole time-of-flight (TOF) mass spectrometry, where multivariate analysis revealed significant differences between the two groups. Several metabolites, including acetylcarnitine, betaine, carnitine, choline, and glycerophosphorylcholine (GPC), were significantly changed. The changes of metabolites were further identified by MALDI-TOF/TOF and IMS. Their spatial distribution and relative quantitation in the kidneys were analyzed by IMS. Carnitine compounds were increased in COR and IM, whereas carnitine and acetylcarnitine were decreased in OM. Choline compounds were increased in COR and OM but decreased in IM from furosemide rats. Betaine and GPC were decreased in OM and IM. Taken together, MALDI-TOF/TOF and IMS successfully provide the spatial distribution and relative quantitation of metabolites in the kidney.

Vasopressin-regulated miRNAs and AQP2-targeting miRNAs in kidney collecting duct cells.

Mature microRNA (miRNA) acts as an important posttranscriptional regulator. We aimed to profile vasopressin-responsive miRNAs in kidney inner medullary collecting duct cells and to identify aquaporin-2 (AQP2)-targeting miRNAs. Microarray chip assay was carried out in inner medullary collecting duct tubule suspensions from rat kidneys in the absence or presence of desmopressin (dDAVP) stimulation (10(-9) M, 2 h). The results demonstrated 19 miRNAs, including both precursor and mature miRNAs, as potential candidates that showed significant changes in expression after dDAVP stimulation (P < 0.05). Nine mature miRNAs exhibiting >1.3-fold changes in expression on the microarray (miR-127, miR-1, miR-873, miR-16, miR-206, miR-678, miR-496, miR-298, and miR-463) were further examined by quantitative real-time PCR, and target genes of the selected miRNAs were predicted. Next, to identify AQP2-targeting miRNAs, in silico analysis was performed. Four miRNAs (miR-32, miR-137, miR-216a, and miR-216b) target the 3'-untranslated region of rat AQP2 mRNA. Target seed regions of miR-32 and miR-137 were also conserved in the 3'-untranslated region of mouse AQP2 mRNA. Quantitative real-time PCR and immunoblot analysis demonstrated that dDAVP-induced AQP2 expression was significantly attenuated in mpkCCDc14 cells when cells were transfected with miRNA mimics of miR-32 or miR-137. Moreover, luciferase reporter assay demonstrated a significant decrease of AQP2 translation in mpkCCDc14 cells transfected with miRNA mimics of miR-32 or miR-137. The present study provides novel insights into the regulation of AQP2 by RNA interference; however, vasopressin-regulated miRNAs did not include miR-32 or miR-137, indicating that the interaction of miRNAs with the AQP2 regulatory pathway requires further analysis.

Uncovering protein polyamination by the spermine-specific antiserum and mass spectrometric analysis.

The polyamines spermidine and spermine, and their precursor putrescine, have been shown to play an important role in cell migration, proliferation, and differentiation. Because of their polycationic property, polyamines are traditionally thought to be involved in DNA replication, gene expression, and protein translation. However, polyamines can also be covalently conjugated to proteins by transglutaminase 2 (TG2). This modification leads to an increase in positive charge in the polyamine-incorporated region which significantly alters the structure of proteins. It is anticipated that protein polyamine conjugation may affect the protein-protein interaction, protein localization, and protein function of the TG2 substrates. In order to investigate the roles of polyamine modification, we synthesized a spermine-conjugated antigen and generated an antiserum against spermine. In vitro TG2-catalyzed spermine incorporation assays were carried out to show that actin, tubulins, heat shock protein 70 and five types of histone proteins were modified with spermine, and modification sites were also identified by liquid chromatography and linear ion trap-orbitrap hybrid mass spectrometry. Subsequent mass spectrometry-based shotgun proteomic analysis also identified 254 polyaminated sites in 233 proteins from the HeLa cell lysate catalyzed by human TG2 with spermine, thus allowing, for the first time, a global appraisal of site-specific protein polyamination. Global analysis of mouse tissues showed that this modification really exists in vivo. Importantly, we have demonstrated that there is a new histone modification, polyamination, in cells. However, the functional significance of histone polyamination demands further investigations.

Patterns of gene and metabolite define the effects of extracellular osmolality on kidney collecting duct.

To investigate the effects of changes in extracellular osmolality on the function of kidney collecting duct cells, particularly on water and sodium reabsorption in the conditions of diuresis and antidiuresis, we generated transcriptome and metabolome profiles of primary cultured inner medullary collecting duct (IMCD) cells. They were grown in hyperosmolar culture medium (640 mOsm) for 4 days and then exposed to either reduced (300 mOsm) or same osmolality for 1 or 2 days more. Integrated analysis of the transcriptome and metabolome revealed that decreased extracellular osmolality was associated with decreased levels of organic osmolytes, glucose, intermediates of citric acid cycle, and branched-chain amino acids (BCAA) in IMCD cells, along with significantly decreased gene expression and protein abundance of P-type transporters (ATP1B1), ABC transporters (ABCC5 and ABCG1), and insulin signaling pathways (IRS2). Quantitative real-time RT-PCR and semiquantitative immunoblotting confirmed the changes of transcript levels of differentially expressed genes and protein levels. Taken together, integrated analysis of omics data demonstrated that water and sodium reabsorption could be reduced by decreased extracellular osmolality per se, through decreased levels of ABC transporters and IRS2, which play a potential role in the transport of organic osmolytes, BCAA, glucose, and trafficking of epithelial sodium channel.

E3 ubiquitin-protein ligases in rat kidney collecting duct: response to vasopressin stimulation and withdrawal.

The E3 ubiquitin (Ub)-protein ligases (E3s) play a role as regulators of protein trafficking and degradation. We aimed to integrate the profile of E3s in rat kidney and examine the changes in protein abundance of the selected E3s in response to 1-deamino-8-D-arginine vasopressin (dDAVP) stimulation/withdrawal. Sprague-Dawley rats were infused with vehicle (n = 13), dDAVP for 5 days (n = 13), or dDAVP was withdrawn for periods (15 min, 30 min, 1, 3, 6, 12, or 24 h) after 5-day infusion (n = 46). Total RNA was isolated from the inner medulla (IM) for transcriptome analysis. Plasma membrane (PM)- or intracellular vesicle (ICV)-enriched fractions of whole kidney were immunoisolated for liquid chromatography-tandem mass spectrometry analysis. dDAVP infusion for 5 days (D5d) significantly increased urine osmolality, which was maintained during 3-h withdrawal of dDAVP after 5-day infusion (D5d-3h). Consistent with this, aquaporin-2 (AQP2) expression in the PM fractions of D5d and D5d-3h increased, whereas AQP2 expression in the ICV fractions of D5d-3h was further increased, indicating internalization of AQP2. Transcriptome analysis revealed 86 genes of E3s and LC-MS/MS analysis demonstrated 16 proteins of E3s. Among these, seven E3s (BRCA1, UBR4, BRE1B, UHRF1, NEDD4, CUL5, and FBX6) were shared. RT-PCR demonstrated mRNA expressions of the seven identified E3s in the kidney, and immunoblotting demonstrated changes in protein abundance of the selected E3s (BRE1B, NEDD4, and CUL5) in response to dDAVP stimulation/withdrawal or lithium-induced nephrogenic diabetes insipidus. The rate of AQP2 degradation was retarded in mpkCCDc14 cells with small interfering RNA-mediated knockdown of NEDD4 or CUL5. Taken together, identified E3s could be involved in the degradation of proteins associated with vasopressin-induced urine concentration.

Emerging role of Akt substrate protein AS160 in the regulation of AQP2 translocation.

AS160, a novel Akt substrate of 160 kDa, contains a Rab GTPase-activating protein (GAP) domain. The present study examined the role of Akt and AS160 in aquaporin-2 (AQP2) trafficking. The main strategy was to examine the changes in AQP2 translocation in response to small interfering RNA (siRNA)-mediated AS160 knockdown in mouse cortical collecting duct cells (M-1 cells and mpkCCDc14 cells). Short-term dDAVP treatment in M-1 cells stimulated phosphorylation of Akt (S473) and AS160, which was also seen in mpkCCDc14 cells. Conversely, the phosphoinositide 3-kinase (PI3K) inhibitor LY 294002 diminished phosphorylation of Akt (S473) and AS160. Moreover, siRNA-mediated Akt1 knockdown was associated with unchanged total AS160 but decreased phospho-AS160 expression, indicating that phosphorylation of AS160 is dependent on PI3K/Akt pathways. siRNA-mediated AS160 knockdown significantly decreased total AS160 and phospho-AS160 expression. Immunocytochemistry revealed that AS160 knockdown in mpkCCDc14 cells was associated with increased AQP2 density in the plasma membrane [135 ± 3% of control mpkCCDc14 cells (n = 65), P < 0.05, n = 64] despite the absence of dDAVP stimulation. Moreover, cell surface biotinylation assays of mpkCCDc14 cells with AS160 knockdown exhibited significantly higher AQP2 expression [150 ± 15% of control mpkCCDc14 cells (n = 3), P < 0.05, n = 3]. Taken together, PI3K/Akt pathways mediate the dDAVP-induced AS160 phosphorylation, and AS160 knockdown is associated with higher AQP2 expression in the plasma membrane. Since AS160 contains a GAP domain leading to a decrease in the active GTP-bound form of AS160 target Rab proteins for vesicle trafficking, decreased expression of AS160 is likely to play a role in the translocation of AQP2 to the plasma membrane.

Novel diagnostic algorithm for the floating and sunken pulse qualities and its clinical test.

We propose a novel classification algorithm for the floating pulse and the sunken pulse using a newly defined coefficient (C(fs)). To examine the validity of the proposed algorithm, we carried out a clinical test in which 12 oriental medical doctors made pairwise diagnoses on the pulses of volunteering subjects. 169 subjects were simultaneously diagnosed by paired doctors, and the diagnoses in 121 subjects were concordant, yielding an accuracy of 72% and a Matthews correlation coefficient of 0.42, which indicates reasonable agreement between doctors. Two sample T-tests showed that subjects in the sunken pulse group had significantly higher BMI and C(fs) (P < .05) than those in the floating pulse group. The pulse classification by the algorithm converged with the diagnoses of paired doctors with an accuracy up to 69%. With these results, we confirm the validity of the novel classification algorithm for the floating and sunken pulses.

Pilot Study to Evaluate the Efficacy of Polynucleotide Sodium Compared to Sodium Hyaluronate and Crosslinked Sodium Hyaluronate in Patients with Knee Osteoarthritis.

Degenerative arthritis of the knee joint has become a major social problem worldwide due to population aging. There are several treatment options for knee osteoarthritis, and the intraarticular injection of sodium hyaluronate is commonly selected by many clinicians as a nonsurgical treatment. However, the efficacy of the treatment is controversial. In this pilot study, we aimed to compare polynucleotide sodium (Conjuran®) with sodium hyaluronate (Hyruan Plus®) and 1,4-butanediol diglycidyl ether-crosslinked sodium hyaluronate (Synovian®) in terms of analgesic efficacy after intraarticular injection in patients with knee osteoarthritis. One of the three intraarticular agents was selected according to what agents were available for outpatients when each patient was enrolled in the study. The 15 enrolled patients were subdivided into 3 groups of 5 patients each. Three injections were performed under ultrasound guidance at a 1-week intervals over a total of 3 weeks. The visual analog scale (VAS) score, the Korean version of the Western Ontario and McMaster Universities Arthritis Index (K-WOMAC), the EuroQol five-dimension scale (EQ-5D) score, and the Korean version of the painDETECT Questionnaire (K-PDQ) score were evaluated before injection and at 1, 2, and 6 weeks after the start of the treatment protocol. The primary endpoint was the change in weight-bearing pain at 4 weeks after the last injection. Secondary endpoints included pain at rest and during walking and the K-WOMAC, EQ-5D, and K-PDQ scores. Weight-bearing pain decreased significantly more from pretreatment to 6 weeks after the start of the treatment protocol in the polynucleotide sodium-treated patients than in the patients who were treated with other agents (p = 0.006, one-way ANOVA). There were no significant between-group differences in the other secondary endpoints. No adverse events occurred. In conclusion, polynucleotide sodium could effectively reduce weight-bearing pain in the patients with knee osteoarthritis compared to standard hyaluronic acid viscosupplementation.

Analgesic Effect of Low Dose Nefopam Hydrochloride after Arthroscopic Rotator Cuff Repair: A Randomized Controlled Trial.

Arthroscopic rotator cuff repair causes acute postoperative hyperalgesia. Multimodal analgesia is preferable to opioid-based intravenous patient-controlled analgesia (IV-PCA) due to postoperative nausea and vomiting (PONV). We evaluated the effect of nefopam as a postoperative non-opioid analgesic after shoulder surgeries. A total of 180 adult patients were enrolled for arthroscopic rotator cuff repair. They were randomly assigned to nefopam (N) or control (C) groups and each group was reclassified according to the interscalene block (B) into NB, CB and NX, CX. Nefopam was applied at a constant dose intravenously during recovery. Pain scores were measured with a Visual Analogue Scale (VAS) before (T0), immediately after (T1), 30 min (T2) and 12 h (T3), 24 h (T4) and 48 h (T5) after surgery. There was no significant difference in demographic data. The overall VAS scores did not differ with regard to nefopam use, except for the NB group at T4 in intention to treat (ITT) analysis (p < 0.05). PONV occurred more frequently in the N group than in the C group (p < 0.05). Neither individual nor all risk factors were associated with PONV occurrence (p > 0.10). In conclusion, nefopam alone did not show a definite decrease in postoperative pain. It instead increased PONV regardless of risk factors.

Short-Term Efficacy of Pulsed Radiofrequency Thermal Stimulation on Acupoints for Chronic Low Back Pain: A Preliminary Study of a Randomized, Single-Blinded, Placebo-Controlled Trial.

BACKGROUND: The objective of this study was to evaluate the pain-relief efficacy of thermal stimulation induced by a pulsed radiofrequency (PRF) thermal stimulation applied to acupoints (APs) in patients with low back pain (LBP). The study was designed as a randomized, single-blinded, placebo-controlled trial. Methods. Fifty-six LBP patients whose minimum pain intensity score on a visual analogue scale (VAS, 0-100 mm) was more than 30 mm were randomly allocated to either the placebo-controlled or the treatment group at a 1:1 ratio. The treatment and placebo-controlled groups received PRF thermal stimulation plus cupping therapy and cupping therapy only, respectively. Each patient was scheduled to receive a total of three treatment sessions over one week with allowing a window up to 4 days. Six of the 13 predefined APs were selected differently for each session depending on the change in patient's symptoms and intensity of pain. The primary outcome was the mean difference between the placebo-controlled and treatment group of VAS changes from the baseline to the end of the follow-up period. RESULTS: The patients' reported VAS scores from baseline to the end of follow-up (average: 9.8 days) were significantly decreased by 8.036 points (two-sided 95% CI, -11.841 to -4.231) and 13.393 points (two-sided 95% CI: 17.198 to -9.588) in the treatment and the placebo-controlled groups, respectively. However, the change in VAS scores between the treatment group and the placebo-controlled group was not significantly different (2.015 mm, two-sided 95% CI: -5.288 to 9.317). CONCLUSION: The trial results indicated that treatment with either PRF thermal stimulation with cupping therapy or cupping therapy alone effectively relieved LBP. The efficacy of PRF thermal stimulation combined with cupping therapy was not superior to that of cupping therapy alone. Trial registration number: Clinical Research Information Service (KCT0002137). The trial was registered retrospectively on 10 November, 2016.

Proposal for a New Predictive Model of Short-Term Mortality After Living Donor Liver Transplantation due to Acute Liver Failure.

BACKGROUND Acute liver failure (ALF) is known to be a rapidly progressive and fatal disease. Various models which could help to estimate the post-transplant outcome for ALF have been developed; however, none of them have been proved to be the definitive predictive model of accuracy. We suggest a new predictive model, and investigated which model has the highest predictive accuracy for the short-term outcome in patients who underwent living donor liver transplantation (LDLT) due to ALF. MATERIAL AND METHODS Data from a total 88 patients were collected retrospectively. King's College Hospital criteria (KCH), Child-Turcotte-Pugh (CTP) classification, and model for end-stage liver disease (MELD) score were calculated. Univariate analysis was performed, and then multivariate statistical adjustment for preoperative variables of ALF prognosis was performed. A new predictive model was developed, called the MELD conjugated serum phosphorus model (MELD-p). The individual diagnostic accuracy and cut-off value of models in predicting 3-month post-transplant mortality were evaluated using the area under the receiver operating characteristic curve (AUC). The difference in AUC between MELD-p and the other models was analyzed. The diagnostic improvement in MELD-p was assessed using the net reclassification improvement (NRI) and integrated discrimination improvement (IDI). RESULTS The MELD-p and MELD scores had high predictive accuracy (AUC >0.9). KCH and serum phosphorus had an acceptable predictive ability (AUC >0.7). The CTP classification failed to show discriminative accuracy in predicting 3-month post-transplant mortality. The difference in AUC between MELD-p and the other models had statistically significant associations with CTP and KCH. The cut-off value of MELD-p was 3.98 for predicting 3-month post-transplant mortality. The NRI was 9.9% and the IDI was 2.9%. CONCLUSIONS MELD-p score can predict 3-month post-transplant mortality better than other scoring systems after LDLT due to ALF. The recommended cut-off value of MELD-p is 3.98.

Differences in the Properties of the Radial Artery between Cun, Guan, Chi, and Nearby Segments Using Ultrasonographic Imaging: A Pilot Study on Arterial Depth, Diameter, and Blood Flow.

Aim of the Study. The three conventional pulse-diagnostic palpation locations (PLs) on both wrists are Cun, Guan, and Chi, and each location reveals different clinical information. To identify anatomical or hemodynamic specificity, we used ultrasonographic imaging to determine the arterial diameter, radial artery depth, and arterial blood flow velocity at the three PLs and at nearby non-PL segments. Methods. We applied an ultrasound scanner to 44 subjects and studied the changes in the arterial diameter and depth as well as in the average/maximum blood flow velocities along the radial artery at three PLs and three non-PLs located more proximally than Chi. Results. All of the measurements at all of the PLs were significantly different (P < 0.01). Artery depth was significantly different among the non-PLs; however, this difference became insignificant after normalization to the arm circumference. Conclusions. Substantial changes in the hemodynamic and anatomical properties of the radial artery around the three PLs were insignificant at the nearby non-PLs segments. This finding may provide a partial explanation for the diagnostic use of "Cun, Guan, and Chi.".

Short-Term Effect of Laser Acupuncture on Lower Back Pain: A Randomized, Placebo-Controlled, Double-Blind Trial.

Purpose. This trial was performed to investigate the efficacy of laser acupuncture for the alleviation of lower back pain. Methods. This was a randomized, placebo-controlled, double-blind trial. Fifty-six participants were randomly assigned to either the laser acupuncture group (n = 28) or the sham laser acupuncture group (n = 28). Participants in both groups received three treatment sessions over the course of one week. Thirteen acupuncture points were selected. The visual analogue scale for pain, pressure pain threshold, Patient Global Impression of Change, and Euro-Quality-of-Life Five Dimensions questionnaire (Korean version) were used to evaluate the effect of laser acupuncture treatment on lower back pain. Results. There were no significant differences in any outcome between the two groups, although the participants in both groups showed a significant improvement in each assessed parameter relative to the baseline values. Conclusion. Although there was no significant difference in outcomes between the two groups, the results suggest that laser acupuncture can provide effective pain alleviation and can be considered an option for relief from lower back pain. Further studies using long-term intervention, a larger sample size, and rigorous methodology are required to clarify the effect of laser acupuncture on lower back pain.

Paenibacillus swuensis sp. nov., a bacterium isolated from soil.

Strain DY6(T), a Gram-positive endospore-forming motile rod-shaped bacterium, was isolated from soil in South Korea and characterized to determine its taxonomic position. Phylogenetic analyses based on the 16S rRNA gene sequence of strain DY6(T) revealed that strain DY6(T) belongs to the genus Paenibacillus in the family Paenibacillaceae in the class Bacilli. The highest degree of sequence similarities of strain DY6(T) were found with Paenibacillus gansuensis B518(T) (97.9%), P. chitinolyticus IFO 15660(T) (95.3%), P. chinjuensis WN9T (94.7%), and P. rigui WPCB173(T) (94.7%). Chemotaxonomic data revealed that the predominant fatty acids were anteiso-C(15:0) (38.7%) and C(16:0) (18.0%). A complex polar lipid profile consisted of major amounts of diphosphatidylglycerol, phosphatidylethanolamine, and phosphatidylglycerol. The predominant respiratory quinone was MK-7. Based on these phylogenetic, chemotaxonomic, and phenotypic data, strain DY6(T) (=KCTC 33026(T) =JCM 18491(T)) should be classified as a type strain of a novel species, for which the name Paenibacillus swuensis sp. nov. is proposed.

Ubiquitination of aquaporin-2 in the kidney.

Ubiquitination is known to be important for endocytosis and lysosomal degradation of aquaporin-2 (AQP2). Ubiquitin (Ub) is covalently attached to the lysine residue of the substrate proteins and activation and attachment of Ub to a target protein is mediated by the action of three enzymes (i.e., E1, E2, and E3). In particular, E3 Ub-protein ligases are known to have substrate specificity. This minireview will discuss the ubiquitination of AQP2 and identification of potential E3 Ub-protein ligases for 1-deamino-8-D-arginine vasopressin (dDAVP)-dependent AQP2 regulation.

A study on characteristics of radial arteries through ultrasonic waves.

The study aims to measure and analyze the thickness and depth of blood vessel on the pulse diagnosis locations (CHON, KWAN, CHUCK) and the blood velocity through the use of ultrasonic waves (LOGIQ5PRO, GE Medical, U.S.) in order to understand the structural difference of pulse diagnosis locations. The subjects included 44 healthy men and women (22.28+/-2.62 age) considered normal in terms of Body Mass Index (BMI). The thickness and depth of the blood vessel and the blood velocity were measured three times on CHON, KWAN and CHUCK to obtain the average value.

A novel method of ligand peptidomics to identify peptide ligands binding to AQP2-expressing plasma membranes and intracellular vesicles of rat kidney.

Aquaporin-2 (AQP2), the vasopressin-regulated water channel in collecting duct principal cells, plays a key role in the regulation of body water balance. We aimed to isolate high-affinity peptide ligands that bind to immunoisolated AQP2-expressing plasma membrane (PM) or intracellular vesicle (ICV) preparations from rat kidney by the in vitro phage display technique. Immunoblotting revealed that AQP2 was exclusively expressed in the immunoisolated AQP2 membrane fractions (PM and ICV), compared with the nonimmunoisolated or preimmune IgG pulldown rat kidney samples. Moreover, AQP1 or H+-ATPase (B1 subunit) expression was minimal in the immunoisolated AQP2 membrane fractions, indicating the specificity of AQP2 membrane isolation. A phage peptide library based on T7 415-1b phage vector displaying CX7C was constructed. After three rounds of biopanning, seven phage clones of high frequency were selected, which showed high affinity to the AQP2-containing PM or ICV fractions compared with a nonrecombinant T7 insertless phage clone. In contrast, these phage clones showed lower affinity to H+-ATPase-containing fractions. Fluorescein-conjugated peptide labeling was associated with intracellular compartment and PM of primary cultured inner medullary collecting duct cells, relative to absent or very weak labeling with fluorescein-conjugated control peptide. Library analyses demonstrated proteins that had motifs homologous to the peptide ligands, albeit with a high probability of a random match due to short peptide sequences. In summary, we applied the in vitro phage display technique to identify high-affinity peptide ligands to AQP2-expressing membranes. Library analyses identified proteins having homologous motifs, which need to be examined for involvement in AQP2 trafficking and regulation.