[Imaging characteristics and surgical effect for symmetrical lumbar hemivertebrae].

Objective: To analyze the imaging characteristics and surgical effect for symmetrical lumbar hemivertebrae in pediatric patients. Methods: The data of 13 patients with hemivertebrae locating in the lumbar spine symmetrically were retrospectively analyzed, and all the patients were treated in Beijing Children's Hospital from January 2015 to September 2021. The mean age of the patients was 6.2 (2.9, 9.3) years. There were 8 males and 5 females. The data of coronal/sagittal plane including segmental Cobb angle, cranial/caudal compensatory curve, thoracic kyphosis, thoracolumbar kyphosis, sacral obliquity, and lumbar lordosis were recorded through long cassette spinal radiographs. Associated anomalies and the relationship between hemivertebrae and posterior component were recorded through computerized tomography (CT) and magnetic resonance imaging (MRI). All the patients received surgery, and their pre-and postoperative imaging data were compared. Results: A total of 26 hemivertebraes were found, in which 80.8% (21/26) located below L2. Hemivertebraes in 10 patients were separated by a mean 1-2 normal vertebrae. Most hemivertebraes along with the corresponding posterior component were unison (21/26, 80.8%). The Cobb angles of cranial compensatory curve (13.9°±7.2°) was more serious than that of caudal compensatory curve (5.5°±5.0°)(P=0.04). The lumbar lordosis and thoracic kyphosis was 20.2°±15.0° and 18.7°±9.2°, respectively. Six patients complicated with sacral obliquity, while 7 patients complicated with thoracolumbar lordosis. Associated anomalies were found in 6 (46.2%) patients through CT and MRI. Eleven patients received one-or two-stage posterior hemivertebrae resection with short segmental fusion, and 2 patients received one-stage hemivertebrae resection with long segmental fusion. All the surgery were completed successfully without serious complications such as nerve injury, infection, and implant failure. The mean follow-up period was (42.4±10.2) months. At the last follow-up point, the correction rate of segmental Cobb angle and cranial compensatory curve was 83.3%±15.6% and 38.1%±10.4%, respectively, showing significant improvement (P<0.05). Although the caudal compensatory curve, sacral obliquity, and thoracic kyphosis improved after surgery, the data showed no significant difference compared to that before surgery. Thoracolumbar lordosis in all patients were corrected. Conclusions: Most hemivertebraes in such spinal deformity locate in lower lumbar region with a high incidence of anomalies. Individualized treatment based on patients' condition is essential for the complicated spinal deformity.

Dual-channel compressed ultrafast photography for Z-pinch dynamic imaging.

The compressed ultrafast photography (CUP) can capture non-repetitive time-evolving events at 7 × 1013 fps, which is anticipated to find a diverse range of applications in physics, biomedical imaging, and materials science. The feasibility of diagnosing ultrafast phenomenon of Z-pinch by using the CUP has been analyzed in this article. Specifically, a dual-channel CUP design has been adopted for acquiring high quality reconstructed images and the strategies of identical masks, uncorrelated masks, and complementary masks have been compared. Furthermore, the image of the first channel was rotated by 90° to balance the spatial resolution between the sweep direction and the non-sweep direction. Both five synthetic videos and two simulated Z-pinch videos were chosen as the ground truth to validate this approach. The average peak signal to noise ratio of the reconstruction results is 50.55 dB for the self-emission visible light video and 32.53 dB for the laser shadowgraph video with unrelated masks (rotated channel 1). The simulation results show that the time-space-evolving process of plasma distribution can be well retold, and the phenomenon of plasma instability can be accurately diagnosed by the dual-channel CUP with unrelated masks (rotated channel 1). This study may promote the practical applications of the CUP in the field of accelerator physics.

[The role of STAT-6/KLF-4/PPAR-γ activation in alveolar macrophage polarization changes in silica-induced pulmonary fibrosis].

Objective: To observe the effect of silicon dioxide (SiO(2)) on the polarization of alveolar macrophages (AMs) , and to explore the expressions and the significance of signal transducer and activator of transcription-6 (STAT-6) /Krüppel-like factor-4 (KLF-4) /peroxisome proliferators-activated receptors-γ (PPAR-γ) signaling molecules in AMs. Methods: In November 2020, C57BL/6 mice were randomly divided into crystalline SiO(2) group and normal saline (NS) group, and 12 mice in each group. Mice were intratracheally instillated with 100 µl crystalline SiO(2) suspension (20 mg/ml) or 100 µl NS, and were sacrificed after 28 days. Masson staining was used to observe the degree of pulmonary fibrosis of mice and hydroxyproline (HYP) level were assessed. The proportions of M1-typed and M2-typed AMs in bronchoalveolar lavage fluid (BLAF) were analyzed by flow cytometry. The mRNA relative expression levels of inducible nitric oxide synthase (iNOS) , arginidase-1 (Arg-1) , interleukin (IL) -1ß, tumor necrosis factor-α (TNF-α) , IL-6, IL-10, transforming growth factor-ß (TGF-ß) , STAT-6, KLF-4 and PPAR-γ were detected by real-time fluorescence quantitative PCR. Activities of iNOS and Arg-1, as well as contents of IL-1ß, TNF-α, IL-6, IL-10 and TGF-ß were assessed by the enzyme-linked immunosorbent. The protein relative expression levels of phosphorylation-signal transducer and activator of transcription-6 (p-STAT-6) , KLF-4 and PPAR-γ were evaluated by immunofluorescence. Results: After 28 days of treatment, the structure of the lung tissue of the mice was destroyed, and the deposition of collagen was significantly increased in the crystalline SiO(2) group. Compared with NS group, HYP level of lung tissue in crystalline SiO(2) group were increased, the proportion of M2-typed AMs in crystalline SiO(2) group was increased, the proportion of M1-typed AMs in crystalline SiO(2) group was decreased, the mRNA relative expressions and contents of Arg-1, IL-10, TGF-ß in crystalline SiO(2) group were significantly increased, the mRNA relative expressions and contents of iNOS, IL-1ß, TNF-α, IL-6 in crystalline SiO(2) group were significantly decreased, the mRNA of STAT-6, KLF-4, PPAR-γ and the protein relative expression levels of p-STAT-6, KLF-4, PPAR-γ were significantly increased in crystalline SiO(2) group, and the the differences were statistically significant (P<0.05) . Conclusion: Crystalline SiO(2) may mediate the process of pulmonary fibrosis through promote AMs polarization toward M2-typed by activating the STAT-6/KLF-4/PPAR-γ signaling pathway.

[The correlation between metabolic parameters in (18)F-FDG PET-CT and solid and micropapillary histological subtypes in lung adenocarcinoma].

Objective: Solid and micropapillary pattern are highly invasive histologic subtypes in lung adenocarcinoma and are associated with poor prognosis while the biopsy sample is not enough for the accurate histological diagnosis. This study aims to assess the correlation and predictive efficacy between metabolic parameters in (18)F-fluorodeoxy glucose positron emission tomography/computed tomography ((18)F-FDG PET-CT), including the maximum SUV (SUV(max)), metabolic tumor volume (MTV), total lesion glycolysis (TLG) and solid and micropapillary histological subtypes in lung adenocarcinoma. Methods: A total of 145 resected lung adenocarcinomas were included. The clinical data and preoperative (18)F-FDG PET-CT data were retrospectively analyzed. Mann-Whitney U test was used for the comparison of the metabolic parameters between solid and micropapillary subtype group and other subtypes group. Receiver operating characteristic (ROC) curve and areas under curve (AUC) were used for evaluating the prediction efficacy of metabolic parameters for solid or micropapillary patterns. Univariate and multivariate analyses were conducted to determine the prediction factors of the presence of solid or micropapillary subtypes. Results: Median SUV(max) and TLG in solid and papillary predominant subtypes group (15.07 and 34.98, respectively) were significantly higher than those in other subtypes predominant group (6.03 and 10.16, respectively, P<0.05). ROC curve revealed that SUV(max) and TLG had good efficacy for prediction of solid and micropapillary predominant subtypes [AUC=0.811(95% CI: 0.715~0.907) and 0.725(95% CI: 0.610~0.840), P<0.05]. Median SUV(max) and TLG in lung adenocarcinoma with the solid or micropapillary patterns (11.58 and 22.81, respectively) were significantly higher than those in tumors without solid and micropapillary patterns (4.27 and 6.33, respectively, P<0.05). ROC curve revealed that SUV(max) and TLG had good efficacy for predicting the presence of solid or micropapillary patterns [AUC=0.757(95% CI: 0.679~0.834) and 0.681(95% CI: 0.595~0.768), P<0.005]. Multivariate logistic analysis showed that the clinical stage (Stage Ⅲ-Ⅳ), SUV(max) ≥10.27 and TLG≥7.12 were the independent predictive factors of the presence of solid or micropapillary patterns (P<0.05). Conclusions: Preoperative SUV(max) and TLG of lung adenocarcinoma have good prediction efficacy for the presence of solid or micropapillary patterns, especially for the solid and micropapillary predominant subtypes and are independent factors of the presence of solid or micropapillary patterns.

A high-speed radiation imaging system based on liquid scintillator filled capillary arrays.

A high-speed radiation imaging system based on an image converter of liquid scintillator filled capillary arrays has been developed, which is sensitive to x rays, gamma rays, and neutrons. This imaging system has advantages of both high spatial resolution and high sensitivity because increasing the thickness of the image converter only leads to little deterioration on imaging resolution. The capillary arrays have dimensions of 150 mm diameter and 50 mm thickness, with 100 µm diameter of each capillary. The fluorescence decay time of the filled liquid scintillator based on the mixture of p-xylene and 2,5-diphenyloxazole has been evaluated to be ∼3 ns with the single photon method under the gamma ray excitation. The spatial resolution has been experimentally evaluated to be about 1.15 and 0.6 mm, under excitation of x rays and neutrons, respectively. The imaging system has been applied for diagnosing the dynamic x-ray spot generated by the rod pinch. Two frames in single shot with 15 ns temporal resolution and 20 ns inter-frame separation time have been obtained, which show the spatiotemporal distribution of the electrons bombarding the tungsten rod, indicating the ability of this imaging system in diagnosing dynamic radiation objects. In addition, the technique of capillary arrays provides a promising path for applications of advanced liquid scintillators in the field of radiation imaging.

[Effect of perioperative accelerated rehabilitation management program for children with congenital spinal deformity].

Objective: To explore the perioperative therapeutic effect of enhanced recovery after surgery (ERAS) in children with congenital spinal deformity and summarize the clinical experience. Methods: Fifty-nine pediatric patients with congenital spinal deformities admitted to Beijing Children's Hospital from May 2020 to January 2021 were included in this study, and all patients underwent posterior spinal osteotomy orthopedic implant fusion with internal fixation. There were 22 males and 37 females, aged (7.4±4.1) years. Patients were divided into ERAS group (n=29) and control group (n=30) according to the management model. Patients in the ERAS group were managed with an accelerated recovery management model during the perioperative period, which mainly included: high protein diet, shortened fasting time, optimized anesthesia protocol, and multimodal analgesia. Patients in the control group received the traditional perioperative management model. The indexes of surgery, diet, pain score and laboratory tests were compared between the two groups. Results: All patients completed the surgery successfully. The mean temperature and pain scores of patients in the ERAS group were lower than those in the control group at 3 days postoperatively (P<0.05). The time to exhaustion and defecation in the ERAS group was (1.0±0.8) d and (2.5±0.9) d postoperatively, both significantly earlier than those in the control group ((3.4±0.8) d and (4.0±1.1) d) (both P<0.05). C-reactive protein was 38(8,46) mg/L in patients of the ERAS group on the day 3 postoperatively, which was significantly lower than that in the control group 47(22,93) mg/L (P=0.023). The hemoglobin level on postoperative day 3 was (110.7±9.6) g/L in the ERAS group, which was significantly higher than that in the control group ((104.5±11.4) g/L) (P=0.029). Postoperative complications occurred in 8(27.6%) and 9(30.0%) patients in the ERAS and control groups, respectively (P=1.000), with mild abdominal pain and bloating being the most common complications in both groups, most of which were not treated specifically. Conclusion: ERAS is a safe and effective perioperative management mode for children with congenital spinal deformity. Compared with the traditional method, it can significantly improve the treatment efficiency and deserve clinical application.

[Detecting of obstructive sleep apnea hypopnea syndrome using a multi-parameter pressure sensitive sleep monitor].

OBJECTIVE: To evaluate the efficacy of an electro-mechanical film-based(EMFi) multi-parameter pressure sensitive sleep monitor(MPSSM)on clinical diagnosis and research significance of obstructive sleep apnea hypopnea syndrome(OSAHS). METHODS: Retrospective analysis was made of 58 test subjects at Peking University Third Hospital with suspected OSAHS who were simultaneously monitored by MPSSM and polysomnography(PSG). The PSG test results were used as the gold standard in evaluating the sensitivity and specificity of OSAHS diagnosis of MPSSM. The test result consistency of sleep apnea and hypopnea index(AHI)and total apnea time of the two methods was evaluated. Real-time waveform comparison of sleep respiratory events of a randomly selected patient diagnosed with OSAHS was performed. RESULTS: For 58 test subjects, 48 were male, 10 were female, with an average age of(40.6±12.2)years. Thirty-nine out of the 58 test subjects were diagnosed with OSHAS by PSG. The sensitivity of MPSSM for OSAHS diagnosis was 92.3%, with 95% confidence interval of 79.1%-98.4%, and the specificity of MPSSM for OSAHS diagnosis was 100%, with 95% confidence interval of 82.3%-100%. Kappa test k=0.887 (P < 0.001) showed OSAHS diagnosis results of the two methods were almost identical. The AHI measured by MPSSM [12.0(2.6-32.2) times/h] and PSG [13.4(3.1-38.8) times/h] were highly correlated (ρ=0.939, P < 0.001). The total apnea time measured by MPSSM [37.9(9.9-80.5) min] and PSG [32.3(8.6-93.0) min] were highly correlated(ρ=0.924, P < 0.001). Bland-Altman plot showed that the consistency between the test results of the two methods was very high. CONCLUSION: As a portable, non-contact, fully automatic monitoring device, MPSSM is reliable in the screening of OSAHS compared with PSG. It is suitable to be promoted and applied in primary medical institutions, nursing homes and domestic usage. However, further research is required in improving the analysis of different sleep phase and the differentiation of central sleep apnea syndrome respiratory events in order to effectively assist medical personnel in making an accurate sleep apnea diagnosis.

[Research advances on assisted analgesic and sedative treatment of burn children].

Burn children would experience pain and anxiety during treatment and rehabilitation, thus analgesic and sedative treatment should be applied throughout the whole process of burn treatment. Due to the unique physiological and psychological status of burn children, analgesic and sedative treatment for burn children is a relatively complex process compared with that for adult burn patients. Therefore, how to provide the effective and safe analgesic and sedative treatment for burn children is a clinical challenge. This paper reviewed the related progress of assisted analgesic and sedative treatment of burn children, in order to provide a reference for clinical treatment.

[Predictors for perioperative blood loss in pediatric patients with congenital scoliosis].

Objective: To identify predictors of massive blood loss after posterior hemivertebra resection for patients with congenital scoliosis. Methods: The data of 106 children with congenital scoliosis were collected from June 2017 to June 2019 in Beijing Children's Hospital. All the cases received posterior hemivertebra resection and internal fixation. The blood volume was estimated by weight and height. The visible blood loss was recorded according to medical record, and the hidden blood loss was calculated by OSTHEO formula. Perioperative information including age, gender, height, weight, Cobb and kyphosis angle, level fused, number of screws, operative time, and laboratory examinations was collected. Then multivariable linear regression was performed to determine the independent risk factors of blood loss. Results: All the surgeries were completed successfully. The mean age of the children was (7.3±2.3) years and the operative time was (162±56) min. The mean fused levels were 3 and the correction rate for deformity was 78.4%. The amount of blood loss was (568±208) ml which accounted for 42.3% of total blood volume. The visible and hidden blood loss was (334±193) ml and (234±199) ml, respectively, and which accounted for 58.8% and 41.2% of total blood loss. Multivariable linear regression analysis indicated that age, preoperative Cobb angle, the time of surgery and the number of fused levels were independent risk factors of total blood loss. Preoperative Cobb angle ≥40°, spinal fusion ≥4 levels and operative duration ≥140 min indicated more blood loss (all P<0.05). Conclusions: The perioperative blood loss of congenital scoliosis is massive with a high percentage of hidden blood loss. The patients with severe deformity, more fused levels and increased operative time brings higher risk of massive blood loss.

[The role of body fat rate in the evaluation of obstructive sleep apnea].

Objective: To investigate the role of body fat ratio in the evaluation of obstructive sleep apnea(OSA). Methods: A retrospective analysis was made on 174 cases (between November, 2017 and April, 2018 showed that) of sleep monitoring in the Department of Otorhinolaryngology in Peking University Third Hospital. The data included the gender, age, body fat rate, body mass index (BMI), neck circumference, and apnea-hypopnea index (AHI). The above data were analyzed by non parametric correlation analysis, receiver operating characterristic (ROC) curve analysis and multiple factor Logistic regression analysis to study the relationship between the gender,age,body fat rate,BMI,neck circumference and other indexes of the patients with AHI. Results: Nonparametric correlation analysis showed that the correlation from strong to weak to AHI among women was BMI (r=0.621, P<0.001),body fat rate (r=0.602, P<0.001), age (r=0.570, P<0.001), neck circumference (r=0.402, P=0.014), respectively. BMI (r=0.599, P<0.001), neck circumference (r=0.493, P<0.001), body fat rate (r=0.318, P<0.001), and age (r=0.256, P=0.003) among men. ROC curve analysis showed that the strong to weak index (area under curve,AUC) of the AHI>15/h among women was the body fat rate (AUC=0.884, P=0.001), BMI(AUC=0.810, P=0.008), neck circumference (AUC=0.759, P=0.027), age (AUC=0.750, P=0.033), and the male was BMI (AUC=0.765,P<0.001), neck circumference (AUC=0.720, P<0.001), age (AUC=0.634, P=0.008), and body fat rate (AUC=0.632, P=0.010), respectively. Multifactor Logistic regression analysis showed that the body fat rate (OR=1.704,95%CI=1.012-2.870) in women was an independent risk factor for AHI greater than 15/h; the age of male (OR=1. 044, 95%CI=1.005-1.085) and BMI (OR=1.285, 95%CI=1.056-1.562) were independent risk factors for AHI greater than 15/h. Conclusion: Body fat rate can be used as a new indicator for predicting the severity of OSA,especially in adult female population. In adult female moderate to severe OSA patients (AHI>15/h), compared with BMI,neck circumference and age,the body fat rate has the greatest correlation with AHI. Compared with BMI,neck circumference and age,the body fat rate has a decisive role in predicting moderate to severe OSA (AHI>15/h).

Surgical treatment of osteoporotic degenerative spinal deformity with expandable pedicle screw fixation: 2-year follow-up clinical study.

BACKGROUND: Osteoporotic bone offers poor purchase for the instrumentation in patients with degenerative spinal deformity (DSD), which could lead to several complications. Recently, augmentation methods to improve pedicle screw fixation have been proposed. This retrospective study was to investigate the clinical and radiographic outcomes of expandable pedicle screws (EPS) in patients with osteoporotic DSD. HYPOTHESIS: Expandable pedicle screws (EPS) provide excellent instrument fixation in patients with osteoporotic DSD, improving radiographic and clinical outcomes. MATERIALS AND METHODS: A total of 27 (6 males and 21 females) DSD patients who underwent orthopedics operation with EPS were retrospectively studied. Full-length standing spinal radiographs were obtained in all patients pre- and postoperatively and again at the two-year follow-up. The functional evaluations before operation and at two-year follow-up were graded with Scoliosis Research Society outcomes instrument-22 (SRS-22) and Oswestry Disability Index (ODI) scoring system. RESULTS: All patients obtained good corrective outcomes on spinal deformity. The preoperative ODI score was 36.7% and reduced to 11.9% at two-year follow-up (p=0.0000). Before operation, the SRS-22 function, pain, appearance and mental scores were 2.7±0.4, 3±0.6, 2.7±0.5 and 2.9±0.6, respectively. The scores at two-year follow-up were significantly improved to 3.8±0.7, 4.2±0.6, 4.3±0.6 and 4.4±0.7, respectively (p=0.0000). The SRS-22 satisfaction score was 4.6±0.4 at two-year follow-up. No instances of screw breakage, loosening or pullout in any patient at follow-up. DISCUSSION: EPS provides excellent instrument fixation in patients with osteoporotic DSD, improving radiographic and clinical outcomes at two years' follow-up. TYPE OF STUDY: Retrospective case series study. LEVEL OF EVIDENCE: IV.

Influence of BOL on hyaluronic acid, laminin and hyperplasia in hepatofibrotic rats.

AIM: To study the anti-hepatofibrosis mechanism of Bie Jia Jian oral liquid (BOL). METHODS: The model was induced by subcutaneous injection of CCl(4). BOL was administered and the change of serum hyaluronic acid (HA) and laminin (LN) was observed and the degeneration of liver cells and the degree of fibre hyperplasia analyzed. Changes of ultra micro-structure in liver cells were observed in some samples. RESULTS: HA was reduced in both the groups with low and high dosage of BOL, which showed a remarkable difference as compared with that of the model group (low dosage group: 376.15 microg/L+/-35.48 microg/L vs 806.07 microg/L+/-98.49 microg/L P<0.05; high dosage group: 340.14 microg/L+/-30.18 microg/L vs 806.07 microg/L+/-98.49 microg/L P<0.05). The LN content of low and high dosage group of BOL was lower than that of model group (low dosage group: 71.99 microg/L+/-8.15 microg/L vs 133.94 microg/L+/-14.45 microg/L P <0.01; high dosage group: 71.68 microg/L+/-11.62 microg/L vs 133.94 microg/L+/-14.45 microg/L P<0.01) and colchicine group (low dosage group: 71.99 microg/L+/-8.15 microg/L vs 118.28 microg/L+/-16.13 microg/L P < 0.05; high dosage group: 71.68 microg/L+/-11.62 microg/L vs 118.28 microg/L+/-16.13 microg/L P <0.05). Examined by Ridit, BOL could reduce the degeneration and necrosis of liver cells (chi(2)=11.99 P<0.05), the degree of fibre hyperplasia (chi(2)=13.24 P<0.05) and the pathological change of ultra micro-structure as well. CONCLUSION: The BOL has certain therapeutic effect on the experiment hepatofibrosis. Its mechanisms might include: protecting the function of liver cells, inhibiting excessive synthesis and secretion of extracellular matrix from hepatic stellate cells, relieving the capillarization of hepatic sinusoid, improving liver micro-circulation, and regulating immune function.

[Microelement-molybdenum and its use for the treatment of children's fluorine-stained teeth].

This paper is a study of the effects of molybdenum on the experimental fluorine poisoning in rats and on sodium molybdate treatment of the F-stain teeth in children. The experimental results show that the high-F and Mo feed animal, the excretion of urine-F was more than high-F, the F-content of teeth was reduced, the pathological change of skeletal tissue was relatively little, when sodium molybdate was used to treat children with fluorosis teeth for 3 months, the hair-F would reduce, whereas the hair-Mo increase the immune function would rise--the PHA skin test positive rate raised from effect 33.3% to 69.2%. It is suggested that Mo can interfere with the F-poisoning effect and that Mo may well be used as z worth medicine for the prevention and control of this illness.

Elimination of apolipoprotein B48 formation in rat hepatoma cell lines transfected with mutant human apolipoprotein B cDNA constructs.

Rat hepatoma McA-RH7777 cell lines transfected with full-length human apolipoprotein (apo) B constructs produce mostly human apoB48 and only small amounts of apoB100, as a result of mRNA editing at codon 2153 (C to U conversion at nucleotide 6666). To abolish the formation of apoB48 and increase the yield of apoB100 and other forms of apoB longer than apoB48, site-specific mutations were introduced at or near the site of apoB mRNA editing. Among four mutations examined, only that in which codon 2153 was converted from CAA (Gln) to CTA (Leu) effectively precluded the formation of apoB48. In this mutant, a stop codon would not be generated even if the C to U conversion occurred. The three other mutations were introduced to disrupt the proposed stem-loop structure encompassing the editing site. Changes made in the third positions of five codons on the 5' side of the edited base or of four codons 3' of the edited base failed to eliminate the production of a protein with the approximate size of apoB48. A construct in which codon 2153 was changed from CAA to GAT (Asp) also failed to eliminate the production of a protein the size of apoB48. Analysis of the region between nucleotides 6200 and 6900 of the cDNA did not detect any prevalent alternate editing sites. Immunoblot analysis using polyclonal antibodies raised against synthetic peptides of human apoB100 indicated that the carboxyl terminus of the apoB48-like proteins probably resides between amino acid residues 2068 and 2129 of apoB100. These results provide some insight into the mechanism of apoB mRNA editing and will facilitate further studies on apoB-containing lipoproteins.

Expression of carboxyl-terminally truncated forms of human apolipoprotein B in rat hepatoma cells. Evidence that the length of apolipoprotein B has a major effect on the buoyant density of the secreted lipoproteins.

We examined the relationship between the size of human apolipoprotein (apo) B and the formation and secretion of apoB-containing lipoprotein particles. Stable transformants of the rat hepatoma cell line McA-RH7777 harboring a variety of human apoB cDNA constructs were established, and these produced carboxyl-terminally truncated apoB proteins (apoB18, -B23, -B28, -B31, -B48, and -B53). Immunoblotting of apoB proteins secreted into the culture medium and fractionated by equilibrium density ultracentrifugation revealed that each of the truncated apoB species was secreted from the cells. The peak densities of the apoB-containing particles decreased as the length of the apoB proteins increased. Apolipoproteins B18 and B23 appeared at the bottom of the salt gradient (d = 1.23 g/ml), whereas particles containing apoB28, -B31, -B37, -B48, and -B53 exhibited progressive decreases in density. The density distribution of secreted apolipoproteins was not affected by the expression or secretion of these recombinant apoB species. As determined by nondenaturing gel electrophoresis, apoB28, -B31, -B37, -B48, and -B53 formed their own discrete particles, and there was a direct correlation between the size of the particles and the length of the apoB species. The efficiency and rate of secretion of these truncated forms of apoB were studied by measuring the decrease of immunoprecipitated 35S-labeled apoB proteins in the cells and their accumulation in the medium. Proteins corresponding to apoB28 or larger were rapidly and efficiently secreted, whereas apoB18 and apoB23 were secreted much more slowly. These data imply that the size of these truncated apoB forms governs the lipid content of the apoB-containing lipoproteins formed as well as the kinetics of secretion.

An expression system for human apolipoprotein B100 in a rat hepatoma cell line.

We have designed an in vitro expression system for human apolipoprotein (apo) B. A full-length human apoB minigene was constructed from cDNA and genomic apoB clones and inserted into a vector where its expression was directed by the cytomegalovirus promoter. The apoB minigene was expressed in a rat hepatoma cell line, McA-RH7777. Human apoB100, which is the ligand for the low density lipoprotein receptor, was secreted in low density lipoprotein or very low density lipoprotein particles, depending on the composition of the medium. A protein with the mobility of apoB48, a structurally related protein involved in cholesterol metabolism, was also produced from the human apoB minigene. This in vitro expression system for human apoB will enable investigators to identify which domains of this protein are involved in processes such as lipoprotein assembly and secretion. This system should also allow investigators to identify definitively the domain in apoB that enables the protein to bind to the low density lipoprotein receptor.

Choline deficiency causes translocation of CTP:phosphocholine cytidylyltransferase from cytosol to endoplasmic reticulum in rat liver.

The choline-deficient rat liver has been chosen as a physiologically relevant model system in which to study the regulation of phosphatidylcholine biosynthesis. When 50-g rats were placed on a choline-deficient diet for 3 days, the activity of CTP:phosphocholine cytidylyltransferase (CT) was increased 2-fold in the microsomes and decreased proportionately in the cytosol. A low titer antibody to CT was obtained from chickens and used to identify the amount of CT protein in cytosol from rat liver. The amount of CT recovered from the choline-deficient cytosol was significantly less than in cytosol from choline-supplemented rats. When hepatocytes were prepared from choline-deficient livers, supplementation of the medium of the cells with choline caused CT to move from the membranes to cytosol within 1-2 h. The activity of another translocatable enzyme of glycerolipid metabolism, phosphatidate phosphohydrolase, was unchanged in cytosol from choline-deficient rat livers, and the microsomal activity of this enzyme was only minimally increased. When the livers were fractionated into endoplasmic reticulum and Golgi, there was a 2-fold increase in the activity on the endoplasmic reticulum from choline-deficient livers but no change in activity associated with Golgi. Thus, the increased association of CT with endoplasmic reticulum in choline-deficient livers appears to be specific to that subcellular fraction, and the subcellular location of other enzymes may not be affected.

Feedback regulation of CTP:phosphocholine cytidylyltransferase translocation between cytosol and endoplasmic reticulum by phosphatidylcholine.

The mechanism for the increased association of CTP:phosphocholine cytidylyltransferase (CT) with membranes of hepatocytes derived from choline-deficient, compared with choline-supplemented rats, has been investigated. The cells were maintained in culture for 4 h in a choline- and methionine-deficient medium. (Methionine is required for synthesis of phosphatidylcholine (PC) via methylation of phosphatidylethanolamine.) Afterward, the cells were incubated +/- choline for various times up to 4 h. In the presence, but not in the absence, of choline there was a translocation of CT activity from membranes to cytosol. During this time period there was no change in the amounts of unesterified fatty acids or diacylglycerol recovered from the hepatocytes. In addition, there was no evidence for a difference in the incorporation of 32P into CT or other cytosolic proteins isolated from hepatocytes +/- choline. In contrast, there was a highly significant correlation between the concentration of PC in the membranes and the increased activity of CT in the cytosol (R = 0.98) and the decreased activity in the membranes (R = 0.93). The concentration of PC could alternatively be altered by incubation of the choline-deficient hepatocytes with methionine or lyso-PC. With either of these supplementations highly significant correlation coefficients were observed between the concentration of PC in membranes and decreased activity of CT in membranes or increased activity in cytosol. The concentration of PC was reduced in the endoplasmic reticulum, but not the Golgi membranes, isolated from choline-deficient compared with choline-supplemented livers. The data suggest that the amount of PC in the endoplasmic reticulum feedback regulates the amount of CT associated with this membrane.

Reduction in VLDL, but not HDL, in plasma of rats deficient in choline.

We have analyzed plasma lipoprotein levels in young male rats fed a choline-deficient diet for 3 days. We confirmed previous studies that choline deficiency promotes 6.5-fold accumulation of triacyglycerol in the liver (23.9 +/- 6.0 versus 3.69 +/- 0.92 mumol/g liver) and reduction of triacylglycerol concentration in plasma by 60% (0.17 +/- 0.04 versus 0.46 +/- 0.10 mumol/mL plasma). Agarose gel electrophoresis showed that the plasma very low density lipoprotein (VLDL) levels were reduced in choline-deficient rats, but the concentration of plasma high density lipoproteins (HDL) was not affected. Sodium dodecyl sulfate - polyacrylamide gel electrophoresis of fractionated plasma lipoproteins revealed that the concentrations of apolipoproteins (apo) BH, BL, and E in VLDL from choline-deficient rats were 37.1, 11.0, and 37.2% of normal levels, respectively. In contrast, the amount of apo A-I, the major one in HDL, was almost unchanged. Correspondingly, there were decreased lipid (mainly phosphatidylcholine and triacylglycerol) levels in VLDL from choline-deficient rats, but no change in the levels of phosphatidylcholine, cholesterol, and cholesterol ester in HDL. There were similar levels of apo B and E (components of VLDL) in homogenates of livers from normal and choline-deficient rats, as determined by immunoblotting. These results support the hypothesis that choline deficiency causes reduction of VLDL, but not HDL, levels in plasma as a consequence of impaired hepatic VLDL secretion.