Protein-truncation mutations in the RP2 gene in a North American cohort of families with X-linked retinitis pigmentosa.
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Amino Acid Sequence↗
Biomedical subjects
D Yan
Publications and source records attributed to D Yan.
Mutations affecting motifs of unknown function in the central domain of nitrogen regulatory protein C.
The positive control function of the bacterial enhancer-binding protein NtrC resides in its central domain, which is highly conserved among activators of sigma54 holoenzyme. Previous studies of a small set of mutant forms specifically defective in transcriptional activation, called NtrC repressor [NtrC(Rep)] proteins, had enabled us to locate various functional determinants in the central domain. In this more comprehensive survey, the DNA encoding a major portion of the central domain was randomly mutagenized and mutated ntrC genes were introduced into the cell via multicopy expression plasmids. DNA sequencing of 95 isolates identified by a preliminary phenotypic screen revealed that the lesions in them caused 55 distinct single amino acid substitutions at 44 different positions. Assays of glnA transcription in vivo and in vitro yielded two conclusions. First, of the 41 mutant proteins that could be purified, 17 (1 known, 16 new) showed no detectable activity in either assay, thus qualifying them as true NtrC(Rep) proteins. These contained residue changes in six of the seven highly conserved regions in the central domain, including two never studied before. Second, some mutant proteins were inactive in vivo but were either marginally or fully active in vitro. Their surprising lack of activity in vivo may be accounted for by high levels of expression, which apparently decreased activation by these mutant proteins but not by wild-type NtrC (NtrCWT). Of particular interest were a subset of these proteins that exhibited greater transcriptional activation than NtrCWT at low concentrations. Their elevated activation capacities remain to be explained.
Amino Acid Sequence↗
Surgical treatment of patent ductus arteriosus (PDA) through mini subaxillary extrapleural approach.
In this study, a new surgical technique for the surgical treatment of patent ductus arteriosus (PDA) is evaluated. The subaxillary extrapleural approach was performed on 836 patients with patent ductus arteriosus. And 20 of these patients were evaluated for postoperative outcomes compared with the routine technique. The results indicated that the new method is safe, less traumatic and has better cosmetic effects. It is concluded in this article that the subaxillary extrapleural approach for PDA is a good modification to the routine surgical technique, that is surgery through the posterolateral approach.
Cardiac Surgical Procedures↗
[Study on the relationship between perinatal hypoxia and concentration of endothelin-1 in amniotic fluid].
OBJECTIVE: To investigate the relationship between endothelin-1 (ET-1) concentration in amniotic fluid (AF) and perinatal hypoxia. METHODS: 161 cases were measured for amniotic fluid(AF) ET-1 levels by radioimmunoassay. 110 cases of normal pregnancy were included in control group among which 30 term pregnancies were simultaneously measured for maternal and umbilical plasma ET-1. 51 cases of intrauterine hypoxia were the study group. RESULTS: (1) The AF ET-1 levels showed increasing trend after 14 weeks (P < 0.01). (2) Fetal plasma ET-1 levels were significantly higher than that of maternal plasma ET-1 levels, but lower than those of AF ET-1 (P < 0.01). The ET-1 levels of umbilical plasma had positive correlation with those of AF ET-1 (r = 0.952, P < 0.01), but there is no correlation with levels of maternal plasma ET-1 (r = 0.338, P < 0.05). (3) In study group, the level of AF ET-1 was elevated with severity of hypoxia, the average level of AF ET-1 in cases of intrauterine hypoxia was (30.654 +/- 5.832) ng/L. In cases of severe neonatal asphyxia it was (960.650 +/- 236.698) ng/L. CONCLUSIONS: The ET-1 exists in AF and gradually increases while pregnancy advanced. AF ET-1 levels can be served as a marker to predict perinatal hypoxia.
Adult↗
[The changes in arteriography in extremities sustaining high tension electric injuries].
OBJECTIVE: Because it is difficult to estimate the extent and degree of damage to the blood vessels of extremities sustaining high tension electrical injuries, arteriography was carried out to evaluate the vascular changes. METHODS: Selective arteriography was carried out 29 times for 26 extremities in 17 patients suffering from > 380 V electrical injuries. RESULTS: The arteriography showed that the injured arteries were narrowed and occluded, and vascular branches decreased or disappeared. Recanalization of the partially occluded arteries or collateral circulation formation was found after decompression. CONCLUSION: Arteriography is valuable to evaluate the vascular changes in high tension electrical injuries of extremities.
Adolescent↗
[Low-tensioned and "b"-like ileocystoplasty].
OBJECTIVE: To perfectly solve the urine-kept and urinary problems of the vesical cancer patients after vesicoectomy. METHODS: We performed low-tensioned and "b"-like ileocystoplasty in 16 patients. RESULTS: After the operation, new vesica urinaria worked well in keeping and emptying urine-urine through the urethra. Residual urine test, chemical test and urography showed that there were no disturbance of body water and electrolyte metetabolism, no reterograde urine in the ureter, no uracratia, and no damage to renal function. CONCLUSIONS: Ileocystoplasty can be used clinically.
Aged↗
[Cardiac metal foreign body: analysis of 21 cases].
OBJECTIVE: To improve early diagnosis and clinical treatment of metal foreign body in the heart. METHODS: In 21 patients, case history, clinical manifestation, chest X-ray film, and echocardiograph were reviewed. 20 patients received operation: emergency exploration (13) and elective operation (7). Four patients were subjected to cardiopulmonary bypass. The other one was not operated on because of absence of symptoms and small foreign body. Foreign bodies mostly bullets, were usually located in the right ventricle. Ventricular fibrillation occurred in 3 patients during operation, two of them were resuscitated. RESULTS: 20 patients but one recovered, a mortality rate of 5%. CONCLUSIONS: Early diagnosis, rapid management, and localization of foreign body are essential to the selection of treatment.
Adolescent↗
[Synthetic assessment of health impact of cerebrovascular diseases on potential health days of life lost].
OBJECTIVE: To assess synthetically the impact on population health caused by cerebrovascular diseases. METHODS: Incidence, duration of disease, disabilities and deaths of cerebrovascular diseases were analyzed with potential health days of life lost (PHDLL) as an indicator in urban areas of Changsha, Hunan Province. RESULTS: The PHDLL caused by cerebrovascular disease totaled 2 624.94 days per thousand of population, 33.01% of them attributed to deaths and 53.44% to chronic disability. CONCLUSION: Hemorrhagic cerebrovascular disease mainly lead to death and ischemic one caused chronic disability.
Adolescent↗
Biochemical characterization and subcellular localization of the mouse retinitis pigmentosa GTPase regulator (mRpgr).
The retinitis pigmentosa GTPase regulator (RPGR) gene encodes a protein homologous to the RCC1 guanine nucleotide exchange factor and is mutated in 20% of patients with X-linked retinitis pigmentosa. We have characterized the full-length and variant cDNAs corresponding to the mouse homolog of the RPGR gene (mRpgr). Comparison with the human cDNA revealed sequence identity primarily in the region of RCC1 homology repeats. As in humans, the mRpgr gene maps within 50 kilobases from the 5'-end of the Otc gene. The mRpgr transcripts are detected as early as E7 during embryonic development and are expressed widely in the adult mice. Variant mRpgr isoforms are generated by alternative splicing and by utilizing two in-frame initiation codons. The products of mRpgr cDNAs migrate aberrantly in SDS-polyacrylamide gels because of a charged domain. In transfected COS cells, the mRpgr protein is isoprenylated and is localized in the Golgi complex. This subcellular distribution is not observed after treatments with brefeldin A or mevastatin and when the conserved isoprenylation sequence (CTIL) at the carboxyl terminus is deleted or mutagenized. These studies suggest a role for the mRpgr protein in Golgi transport and form the basis for investigating the mechanism of photoreceptor degeneration in X-linked retinitis pigmentosa.
Alternative Splicing↗
Mutation of a gene encoding a protein with extracellular matrix motifs in Usher syndrome type IIa.
Usher syndrome type IIa (OMIM 276901), an autosomal recessive disorder characterized by moderate to severe sensorineural hearing loss and progressive retinitis pigmentosa, maps to the long arm of human chromosome 1q41 between markers AFM268ZD1 and AFM144XF2. Three biologically important mutations in Usher syndrome type IIa patients were identified in a gene (USH2A) isolated from this critical region. The USH2A gene encodes a protein with a predicted size of 171.5 kilodaltons that has laminin epidermal growth factor and fibronectin type III motifs; these motifs are most commonly observed in proteins comprising components of the basal lamina and extracellular matrixes and in cell adhesion molecules.
Amino Acid Sequence↗
Ammonia acquisition in enteric bacteria: physiological role of the ammonium/methylammonium transport B (AmtB) protein.
Homologues of the amtB gene of enteric bacteria exist in all three domains of life. Although their products are required for transport of the ammonium analogue methylammonium in washed cells, only in Saccharomyces cerevisiae have they been shown to be necessary for growth at low NH4+ concentrations. We now demonstrate that an amtB strain of Escherichia coli also grows slowly at low NH4+ concentrations in batch culture, but only at pH values below 7. In addition, we find that the growth defect of an S. cerevisiae triple-mutant strain lacking the function of three homologues of the ammonium/methylammonium transport B (AmtB) protein [called methylammonium/ammonium permeases (MEP)] that was observed at pH 6.1 is relieved at pH 7.1. These results provide direct evidence that AmtB participates in acquisition of NH4+/NH3 in bacteria as well as eucarya. Because NH3 is the species limiting at low pH for a given total concentration of NH4+ + NH3, results with both organisms indicate that AmtB/MEP proteins function in acquisition of the uncharged form. We confirmed that accumulation of [14C]methylammonium depends on its conversion to gamma-N-methylglutamine, an energy-requiring reaction catalyzed by glutamine synthetase, and found that at pH 7, constitutive expression of AmtB did not relieve the growth defects of a mutant strain of Salmonella typhimurium that appears to require a high internal concentration of NH4+/NH3. Hence, contrary to previous views, we propose that AmtB/MEP proteins increase the rate of equilibration of the uncharged species, NH3, across the cytoplasmic membrane rather than actively transporting-that is, concentrating-the charged species, NH4+.
Bacterial Proteins↗
The use of adaptive radiation therapy to reduce setup error: a prospective clinical study.
PURPOSE: Adaptive Radiation Therapy (ART) is a feedback treatment process that optimizes a patient's treatment according to the patient specific information measured during the course of treatment. Utilizing an electronic portal imaging device (EPID) and a computer-controlled multileaf collimator (MLC), the ART process is currently being implemented in our clinic to improve the treatment accuracy by compensating for the treatment setup error. A prospective study was conducted to evaluate the feasibility and efficacy of the ART process for clinical use. METHODS AND MATERIALS: The prospective study included 20 patients who underwent conventional radiotherapy on a linear accelerator equipped with an EPID and a MLC. No specific changes were made in the routine clinical procedures except daily portal images were obtained for each treatment field. Two-dimensional setup error for each treatment field was then measured offline using a software tool. The measured setup errors from initial treatment days were used to predict the systematic and random setup errors for each treatment field. An adjustment decision was made if the predicted systematic error was larger than or equal to 2 mm. Furthermore, the treatment field was extended if the predicted random setup error could not be effectively compensated by the predefined treatment setup margin. Instead of the conventional approach of patient repositioning, setup adjustment was implemented by reshaping the MLC field. The entire process from measuring setup error to reshaping the MLC field was performed offline through a computer network. After completion of a patient's treatment, the systematic and random setup errors after adjustment were compared with those predicted prior to the adjustment. The accuracy of the adjustment, and the reliability and stability of the process were analyzed. RESULTS: Treatment fields of 13 patients were modified to correct for systematic errors. The mean systematic error was 4 mm with a range of 2 to 7 mm before adjustment. It was reduced to 0.5 mm with a range of 0.2 to 1.4 mm after adjustment. There was no significant difference in random setup errors before and after adjustment. The ART process was found to be stable, as more than 95% of patient specific setup margins were predictable within 1 mm using the first four to nine fractions of treatment, confirming the feasibility of treatment plan reoptimization with the ART process. CONCLUSIONS: The prospective study demonstrates that the ART process can be effectively implemented in routine clinical practice to improve treatment accuracy. This process is also ready to be further extended to reoptimize the treatment plan by incorporating the predicted patient specific setup variation.
Feasibility Studies↗
Spatial reproducibility of the ring and tandem high-dose rate cervix applicator.
PURPOSE: High-dose rate (HDR) applicators for uterine cervix brachytherapy are used with increasing frequency. Because multiple HDR fractions are required for treatment, the applicator position reproducibility is of most importance. To study this effect, the clinical data from patients with uterine cervix cancer were examined retrospectively to evaluate the interfraction geometric variation of the HDR applicator and its potential treatment impact. METHODS AND MATERIALS: Eighteen patients with invasive cervical cancer who were treated with definitive radiotherapy at William Beaumont Hospital were included in the study. Patients were treated with 45-50.4 Gy megavoltage external beam to the pelvis, and 35 Gy to the prescription point A from 7 fractions of HDR brachytherapy. The 3-dimensional (3D) interfraction geometrical variation of the ring and tandem (R & T) applicator was measured using predefined reference points in the 7 sets of orthogonal simulation films obtained prior to each HDR application. Spatial reproducibility of the R & T insertion and time-trend of the R & T position variation related to patient's anatomy during the treatment course were analyzed with respect to different groups of patients who had either early or advanced disease. RESULTS: The translational variation of the applicator position for all patients was 6.5, 5.9, and 7.7 mm (one standard deviation), respectively, in the patient's superior-to-inferior (SI), right-to-left lateral (RL), and anterior-to-posterior (AP) direction. The rotational variation was 3.4, 4.6, and 6.0 degrees (one standard deviation) in the patient's coronal, transverse, and sagittal planes. When the patients were grouped based on early disease or advanced disease, the latter demonstrated substantially larger variation (factor of 2) in the applicator position than the former. Furthermore, the time-trend of position variation was observable for both groups of patients. The variations occurred primarily during the first 3 fractions. CONCLUSIONS: Based on the good spatial reproducibility observed in our study, the current clinical procedure for the HDR R & T applicator placement is reliable. Positional reproducibility of the R & T applicator is highly dependent upon the size of tumor volume, which, in turn, deviates the applicator during the early course of HDR brachytherapy. Attention to the construction of the midline block is of paramount importance.
Brachytherapy↗
Role of macrophages in elevated IgA and IL-6 production by Peyer's patch cultures following acute oral vomitoxin exposure.
Oral vomitoxin (VT) exposure in mice results in elevated cytokine gene expression, increased production of IgA, and IgA nephropathy. To determine the potential role of macrophages (Mphi) in these effects, an ex vivo model was devised whereby Peyer's patch (PP) and spleen cells were prepared from mice 2 h after oral exposure to 0 or 25 mg/kg body wt VT, cultured, and then evaluated for IgA and cytokine IL-6 production. Both PP and, to a lesser extent, spleen cells from treatment mice produced more IgA over a 7-day period than did corresponding control cells when cultured without a costimulus or in the presence of either phorbol myristate acetate plus ionomycin (PMA + ION) or lipopolysaccharide (LPS); IgA elevation was most marked in LPS-treated cultures. The VT effect was completely ablated in PP cultures that were depleted of Mphi but not in Mphi-depleted spleen cultures. VT exposure similarly increased production of IL-6, an important helper factor for IgA secretion, in LPS-stimulated PP and spleen cell cultures. IL-6 production was also ablated by Mphi depletion. A potential costimulatory role for Mphi was further suggested because both IgA and IL-6 production increased when Mphi-depleted PP cells from VT-treated animals were cocultured with peritoneal Mphi from VT-treated animals. Similar effects were observed when an analogous ex vivo approach was used with purified PP B cells and peritoneal Mphi. PP B cells from control animals also secreted elevated levels of IgA when cocultured with splenic CD4(+) cells from VT-treated animals, thus confirming previous studies showing that T cell help also contributes to increased IgA production. Potential roles for soluble mediators and cell contact in this process were suggested when IgA production was measured in cultures of PP cells separated from VT-treated Mphi by a semipermeable membrane. Taken together, these and previous results suggest that Mphi may play a key mechanistic role in elevated IgA production and IgA nephropathy in VT-exposed mice.
Administration, Oral↗
Induction of cytokine gene expression in mice after repeated and subchronic oral exposure to vomitoxin (Deoxynivalenol): differential toxin-induced hyporesponsiveness and recovery.
A single oral exposure to vomitoxin (VT) in mice has been previously shown to induce in lymphoid tissues the rapid expression of cytokine mRNAs that are produced by both macrophages and T cells. To determine whether prior VT exposures positively or negatively modulate the cytokine response to the toxin in this model, we evaluated the effects of short-term oral (two to seven consecutive daily doses) and subchronic dietary (4 weeks) exposure to VT on expression of a panel of cytokine mRNAs. Effects of a single oral exposure to 0, 5, and 25 mg/kg body wt of VT or of two such daily consecutive doses on splenic cytokine mRNA abundance were compared 2 h after the last toxin administration using RT-PCR in combination with hybridization analysis. While robust cytokine mRNA responses occurred after a single VT exposure, attenuated but significant induction of interleukin (IL)-6, tumor necrosis factor (TNF)-alpha, IL-1beta, and IL-12p40 mRNA was observed after a second VT dose. Similar but insignificant trends occurred with interferon (IFN)-gamma, IL-2, IL-4, and IL-10 mRNAs. Serum TNF-alpha and IL-6 proteins mimicked cytokine mRNA responses although attenuation responses were less marked. Mice were also dosed with VT at 0, 0.5, 2, or 5 mg/kg body wt consecutively for 2, 4, or 7 days and cytokine mRNAs were assessed 2 h after the last treatment in spleen and Peyer's patches. Upon exposure to 2 and 5 mg/kg body wt VT, the relative abundance of IL-1beta, IL-6, TNF-alpha, IL-12 p35, IL-12p40, IL-2, and IL-10 mRNAs increased with dose frequency whereas IFN-gamma and IL-4 mRNAs were unaffected. When mice were fed 0, 10, and 25 ppm VT for 4 weeks, increased expression of mRNAs for TNF-alpha, IL-2, IFN-gamma, and IL-10 was most prominent. However, when VT-fed mice were also challenged with an oral dose of VT equivalent to daily intake at 2 h prior to RNA isolation, vigorous mRNA responses were observed for IL-1beta, IL-6, TNF-alpha, IL-12p40, IL-12p35, IL-2, IFN-gamma, IL-4, and IL-10. In general, spleens were more responsive to the above effects than Peyer's patches. The results indicate that, following a single prior VT exposure, a significant but attenuated cytokine mRNA response occurred upon a second VT treatment. This hyporesponsiveness was overcome upon repeated exposures to the toxin. These data further support the contention that elevated cytokine expression may play a contributory role in the pathophysiologic and immunologic effects of VT and other trichothecene mycotoxins.
Administration, Oral↗
Interaction of D-tubocurarine analogs with the 5HT3 receptor.
D-Tubocurarine is a potent competitive antagonist of two members of the ligand-gated ion channel family, the muscle-type nicotinic acetylcholine receptor (AChR) and serotonin type-3 receptor (5HT3R). We have used a series of analogs of D-tubocurarine to determine the effects of methylation, stereoisomerization and halogenation on the interaction of D-tubocurarine with the 5HT3R. The affinities of the analogs for the 5HT3R span a 200-fold concentration range and fall into three broad groups. The first group, with affinity constants (Ki) < 150 nM, consists of D-tubocurarine and analogs modified at the nitrogens or 7' hydroxyl. The fact that these compounds all have high affinity for the 5HT3R suggests that these portions of the ligand do not make interactions with the receptor that are critical for high-affinity binding. The second group, with Ki's in the 1-5 microM range, consists of analogs modified at the 12'-hydroxyl or the adjacent 13'-carbon, which suggests that this portion of the ligand makes interactions that are important for high-affinity binding. The third, very low affinity, group is a compound with altered stereoconfiguration at the 1 carbon, demonstrating the importance of proper configuration of the antagonist in ligand-receptor interactions. For the most part, this pattern of selectivity is similar to that for the AChR, suggesting that the structures of the ligand-binding sites of these two receptors share common structural features.
Animals↗
Experimental murine IgA nephropathy following passive administration of vomitoxin-induced IgA monoclonal antibodies.
Oral exposure of mice to vomitoxin (VT) induces elevated levels of serum IgA, circulating IgA immune complexes (IgA-IC), mesangial IgA deposition and haematuria, which all mimic the clinical signs of human IgA nephropathy (IgAN). To further assess the effects of VT-induced IgA in the murine model, B6C3F1 and BALB/C mice were injected intraperitoneally with affinity-purified monoclonal IgA derived from Peyer's patch hybridomas of VT-exposed mice. In B6C3F1 mice, serum IgA, IgM and IgA-IC levels were increased two- to fivefold in treatment groups after 4 and 6 wk compared with controls, whereas increases in serum IgG as high as 18-fold were observed. Urinary erythrocyte counts were also significantly elevated in treatment groups after 2, 4 and 6 wk compared with controls. Concurrent increases in IgA and IgG complexes containing casein, the dietary protein source, occurred in treatment mice. Mesangial IgA, IgG, IgM and C3 deposition were significantly increased in all treatment mice after 6 wk. Electron-dense deposits occurred in the glomeruli of IgA-injected mice after 6 wk. All the above parameters were similarly affected in BALB/C mice. Injection of IgA-secreting hybridoma cells into BALB/C mice increased serum IgA, IgA-IC and IgG levels as well as elevated mesangial IgA, IgG and C3 deposition and haematuria after 2-3 weeks compared with controls. In total, these data indicate that passive administration of VT-induced IgAs can induce the hallmarks of IgA nephropathy. Casein, an antigen found in the diet used for these mice, appeared to form IC with IgA or IgG and these IC may participate in the pathogenesis of this nephropathy.
Animals↗
CUS2, a yeast homolog of human Tat-SF1, rescues function of misfolded U2 through an unusual RNA recognition motif.
A screen for suppressors of a U2 snRNA mutation identified CUS2, an atypical member of the RNA recognition motif (RRM) family of RNA binding proteins. CUS2 protein is associated with U2 RNA in splicing extracts and interacts with PRP11, a subunit of the conserved splicing factor SF3a. Absence of CUS2 renders certain U2 RNA folding mutants lethal, arguing that a normal activity of CUS2 is to help refold U2 into a structure favorable for its binding to SF3b and SF3a prior to spliceosome assembly. Both CUS2 function in vivo and the in vitro RNA binding activity of CUS2 are disrupted by mutation of the first RRM, suggesting that rescue of misfolded U2 involves the direct binding of CUS2. Human Tat-SF1, reported to stimulate Tat-specific, transactivating region-dependent human immunodeficiency virus transcription in vitro, is structurally similar to CUS2. Anti-Tat-SF1 antibodies coimmunoprecipitate SF3a66 (SAP62), the human homolog of PRP11, suggesting that Tat-SF1 has a parallel function in splicing in human cells.
Amino Acid Sequence↗