Predicting the response to neoadjuvant chemoradiation for rectal cancer using nomograms based on MRI tumour regression grade.

PURPOSE: This study aimed to develop nomograms that combine clinical factors and MRI tumour regression grade to predict the pathological response of mid-low locally advanced rectal cancer to neoadjuvant chemoradiotherapy. METHODS: The retrospective study included 204 patients who underwent neoadjuvant chemoradiotherapy and surgery between January 2013 and December 2021. Based on pathological tumour regression grade, patients were categorized into four groups: complete pathological response (pCR, n=45), non-complete pathological response (non-pCR; n=159), good pathological response (pGR, n=119), and non-good pathological response (non-pGR, n=85). The patients were divided into a training set and a validation set in a 7:3 ratio. Based on the results of univariate and multivariate analyses in the training set, two nomograms were respectively constructed to predict complete and good pathological responses. Subsequently, these predictive models underwent validation in the independent validation set. The prognostic performances of the models were evaluated using the area under the curve (AUC). RESULTS: The nomogram predicting complete pathological response incorporates tumour length, post-treatment mesorectal fascia involvement, white blood cell count, and MRI tumour regression grade. It yielded an AUC of 0.787 in the training set and 0.716 in the validation set, surpassing the performance of the model relying solely on MRI tumour regression grade (AUCs of 0.649 and 0.530, respectively). Similarly, the nomogram predicting good pathological response includes the distance of the tumour's lower border from the anal verge, post-treatment mesorectal fascia involvement, platelet/lymphocyte ratio, and MRI tumour regression grade. It achieved an AUC of 0.754 in the training set and 0.719 in the validation set, outperforming the model using MRI tumour regression grade alone (AUCs of 0.629 and 0.638, respectively). CONCLUSIONS: Nomograms combining MRI tumour regression grade with clinical factors may be useful for predicting pathological response of mid-low locally advanced rectal cancer to neoadjuvant chemoradiotherapy. The proposed models could be applied in clinical practice after validation in large samples.

[Study on the diet balance index and its relationship with blood uric acid of smoking adults in Tianjin].

Objective: Diet balance index (DBI_16) was used to evaluate the dietary status of smoking adults in Tianjin, and the relationship between DBI_16 and serum uric acid was analyzed. Methods: A total of 1 478 inhabitants aged 18 and above were enrolled. The dietary status was obtained with a 3-day dietary recall and condiment weighing method. And their fasting venous blood was collected to detect uric acid. Food intake and DBI_16 scores of smokers and non-smokers were described, and the relationship among smoking, diet quality distance (DQD-DBI), and serum uric acid was analyzed. Results: The intake of vegetables, fruits, and milk of smokers in Tianjin was lower than while the salt, oil, cereals, and aquatic products were higher than that of non-smokers. The DBI_16 scores of vegetables and fruits, food types, milk, and beans of smokers were lower than those of non-smokers in Tianjin, and the scores of pure energy foods and condiments were higher than those of the non-smokers. The DQD-DBI, high bound score and low bound score of smokers in Tianjin were 42.0, 14.0, and 29.0 respectively, which were all higher than those of non-smokers. The main problems appeared as moderately inadequate intake (accounting for 67.0%), low, excessive intake (accounting for 70.9%), and moderate imbalance of intake (accounting for 67.2%). The serum uric acid of smokers was higher than in the non-smokers, and there the same result appeared under the conditions of "not suitable" in DQD-DBI. Conclusions: In Tianjin, the dietary imbalance was more evident in smokers than the non-smokers, and the serum uric acid was significantly higher than the non-smokers. Strategies as strengthening the nutrition education and intervention targeted for smokers were in urgent need.

[Study of influence of birth weight on risk of chronic diseases and blood uric acid in adulthood].

Objective: To understand the influence of birth weight on the risk of chronic diseases, such as hypertension and diabetes, and the relationship between birth weight and serum uric acid in adulthood. Methods: According to the Chinese Adult Chronic Disease and Nutrition Surveillance Program, a total of 1 131 residents aged 18 years and above were enrolled from Hexi, Nankai, Hongqiao, Wuqing, Jinnan, Baodi and Jizhou districts of Tianjin. The data of birth weight and blood pressure of the residents were collected, and fasting venous blood samples were collected from them to detect uric acid, blood glucose and blood lipids levels. The distribution of birth weight of the surveyed population was described, and the relationship between birth weight and chronic diseases, such as hypertension and diabetes, and the blood uric acid level in adulthood were analyzed. Results: The average birth weight of the surveyed population in Tianjin was 3.37 kg, which was higher in males than in females. No matter overweight/obesity, hypertension or diabetes, the prevalence rate of normal birth weight was the lowest in adulthood. After univariate logistic regression analysis and adjusting for age, sex, smoking and drinking status, it was found that compared with normal birth weight, low birth weight had a stronger correlation with diabetes (OR=2.91,95%CI:1.46-5.76) and dyslipidemia (OR=1.79,95%CI:1.01-3.19) in adulthood. Macrosomia was strongly associated with overweight/obesity in adulthood (OR=1.47, 95%CI:1.08-2.01). There was no significant difference in serum uric acid level among the residents with different birth weights. Conclusions: The low birth weight of the surveyed population in Tianjin was closely related to the risk of diabetes and dyslipidemia in adulthood, and the macrosomia was closely related to the risk of overweight/obesity in adulthood.

[Study on the effect of slightly acid hypochlorous water on dental unit waterlines biofilm model].

Pseudomonas aeruginosa biofilm was established on PU and PVC's surface by Center of Disease Control (CDC) Biofilm Reactor, to evaluate the effectiveness of Slightly Acidic Hypochlorous water (SAHW) for removing Pseudomonas aeruginosa biofilm on Polyurethane (PU) or Polyvinyl Chloride (PVC), which are commonly used in dental unit waterlines (DUWL). The biofilms were treated with 10 mg/L SAHW, 40 mg/L SAHW and sterile distilled water (control group) by the continuous immersion method for 3, 7, and 10 days. The total viable count (TVC) and Confocal laser scanning microscopy (CLSM) were selected to evaluate the biofilm removal effect and calculate the biofilm reduction rate. The results of TVC showed that 99%-100% biofilm reduction could be achieved in 10 mg/L SAHW group and 40 mg/L SAHW group. The results of CLSM showed that 10 mg/L SAHW group and 40 mg/L SAHW group could reach 89%-100% biofilm reduction after 10 days of treatment.

[Anatomical recognition of pancreatic membrane and its application in surgery].

With the rise of domestic membrane anatomy and preliminary establishment of theoretical framework, the operation concepts supported by membrane anatomy are gaining popularity in surgery, especially in abdominal surgery. However, on account of a deep location and the complexity of organs and tissues around the pancreas and mesangial membrane, there is no unified understanding about the pancreas mesangial by experts and scholars. Meanwhile, few studies on it have been conducted. In addition, the location and extent of total mesangectomy based on the mesangial pancreatic theory are also controversial. The purpose of this article is to summarize the anatomy of pancreatic membrane and its application in surgery, in order to provide support for current studies on pancreatic mesangial anatomy.

[Relationship between both duration and quality of sleep, and metabolic syndrome identified by classification tree model in midlife population in Tianjin].

Objective: To study the association of duration and quality of sleep with metabolic syndrome. Methods: Multi-stage cluster random sampling method was used to extract data from the '2015 adult chronic disease and nutrition monitoring population project' from 42 communities of 7 districts in Tianjin. A total of 1 388 residents aged 45 to 59 years old were selected to analyze the relationship between both duration and quality of sleep and the metabolic syndrome. χ(2) test, non-conditional logistic regression and classification tree models were used for data analysis. Result: The prevalence of metabolic syndrome appeared as 30.4%, 37.4 % and 43.1% in the poor, common or well sleep groups respectively. The prevalence rates of metabolic syndrome in the sleep time <6.0 h and >7.5 h, 6.0-7.5 h groups were 36.5% and 30.6% respectively. After adjusting for gender and age, results from the logistic regression analysis showed that short or long sleep duration, as well as poor sleep quality all appeared as risk factors on metabolic syndrome (P<0.05). According to the classification tree model, factors as gender (importance: 0.004, standardized importance: 100%), quality of sleep (importance: 0.004, standardized importance: 99.5%), duration of sleep (importance: 0.002, standardized importance: 38.6%), education level (importance: 0.001, standardized importance: 22.3%) and salt intake (importance: 0.001, standardized importance: 22.2%) were all important on metabolic syndrome and with interactive effects. Conclusions: Both quality and duration of sleep were important influencing factors on metabolic syndrome among midlife population in Tianjin. More attention should be paid to sleep and health status among the midlife population. Ability on self-management of health should also be strengthened through health education, to prevent metabolic syndrome and other chronic diseases in this population.

[Dietary intake of sodium and potassium among adult inhabitants and evaluation of survey methods in Tianjin in 2015].

Objective: To understand the status and food sources of daily dietary intake of sodium and potassium among adult inhabitants in Tianjin city, and to compare the consistence of sodium and potassium intake determined by combination of 3 consecutive 24-hour recalls and weighing family condiments (3 d dietary survey method) with 24 h urine sodium and potassium determination. Methods: Multistage cluster sampling method was applied to select 1 955 adult inhabitants from 840 households of 7 districts in Tianjin in 2015. Questionnaire survey was used to obtain basic information, 3 d dietary method was used to obtain dietary information, and 450 subjects was selected by systematic random sampling method for 24 h urine collection and determination of sodium and potassium, and 153 complete 24 h urine samples were selected by the coefficient of urinary creatinine excretion, then the consistence of the determination was compared with 3 d dietary survey method. Results: 1 828 subjects were enrolled in this study with age of (54.6±14.7) years, among which 858 were male (46.9%). The average standard intake of sodium and potassium per people per day were (5 631.2±3 372.1) mg and (1 662.1±750.5) mg, while the average actual daily intake were (5 315.8±3 196.5) mg and (1 560.7±696.8) mg. 98.7% (1 804/1 828) of the subjects had dietary sodium intake exceeding the recommended adequate intake of China residents(1 300-1 500 mg/d), while 78.5% (1 435/1 828) consumed dietary potassium below the recommended adequate intake(2 000 mg/d). The main resource of sodium was condiments, among which cooking salt accounted for 61.5%, soy sauce accounted for 11.2%. Potassium mainly came from cereals (32.4%) and vegetables (18.1%). Compared with the 24 h urine sodium method, 3 d dietary method overestimated the sodium intake with a gap [median (P(25), P(75))] as 401.7 (-1 130.6, 1 939.5) mg/d with statistical difference (P<0.05). The gap [median (P(25)-P(75))] for potassium between the two methods was 79.1 (-577.5, 565.2) mg/d, without statistical difference (P>0.05). Conclusion: The dietary intake of sodium is high while potassium is low among adults inhabitants in Tianjin city. Compared with the 24 h urine sodium and potassium determination, the 3 d dietary method has a good consistency in evaluating the dietary potassium intake level of the population, but overestimates the dietary sodium intake.

Favorable effects of hydroxychloroquine on serum low density lipid in patients with systemic lupus erythematosus: A systematic review and meta-analysis.

AIMS: Hydroxychloroquine (HCQ) has shown to have significant immunomodulatory effects in the treatment of systemic lupus erythematosus (SLE). Current studies show favorable effects of HCQ on traditional cardiac risk factors in patients with SLE. This review examined the effects of HCQ on serum low-density lipoprotein (LDL) level in patients with SLE. METHODS: A systematic search of seven major literature search databases from their inception until 3 April, 2017 identified nine studies. Random-effects pooled mean difference with corresponding 95% confidence intervals (CI) were estimated. Heterogeneity was measured by I2 . Publication bias was assessed by visual inspection of funnel plots. Sensitivity analysis examined whether HCQ effect on serum total cholesterol level was similar to the main analysis. The Grading of Recommendations Assessment, Development, and Evaluation system was used to assess the overall quality of evidence. RESULTS: Pooled study participants were 559 patients from eight observation studies (two before-after studies; six case-control studies) examining the effects of HCQ on serum LDL. Pooled study participants' characteristics were as follows: mean age 45.719, female 95.262%, and prednisone use 58.366%. HCQ reduced mean LDL levels by 24.397 mg/dL (95% CI 8.921-39.872; P = 0.002). The number of studies identifying statin use was too few to perform meta-regression analysis of statin use. Heterogeneity was extensive (I2 = 94.739%). Symmetrical funnel plot visualized no evidence of publication bias. CONCLUSION: HCQ was associated with serum LDL level reduction by mean 24.397 mg/dL in patients with SLE. Future prospective studies are need to fully characterize the treatment effect.

Magnesium-Doped Li1.2[Co0.13Ni0.13Mn0.54]O2 for Lithium-Ion Battery Cathode with Enhanced Cycling Stability and Rate Capability.

Mg-doped Li[Li0.2-2xMgxCo0.13Ni0.13Mn0.54]O2 is synthesized by introducing Mg ions into the transition-metal (TM) layer of this layered compound for substituting Li ions through a simple polymer-pyrolysis method. The structural and morphological characterization reveals that the doped Mg ions are uniformly distributed in the bulk lattice, showing an insignificant impact on the layered structure. Electrochemical experiments reveal that, at a Mg doping of 4%, the Li[Li0.16Mg0.04Co0.13Ni0.13Mn0.54]O2 electrode can deliver a larger initial reversible capacity of 272 mAh g(-1), an improved rate capability with 114 mAh g(-1) at 8 C, and an excellent cycling stability with 93.3% capacity retention after 300 cycles. The superior electrochemical performances of the Mg-doped material are possibly due to the enhancement of the structural stability by substitution of Li by Mg in the TM layer, which effectively suppresses the cation mixing arrangement, leading to the alleviation of the phase change during lithium-ion insertion and extraction.

Improved Electrochemical Performance of Fe-Substituted NaNi0.5Mn0.5O2 Cathode Materials for Sodium-Ion Batteries.

A series of O3-phase NaFe(x)(Ni0.5Mn0.5)(1-x)O2 (x = 0, 0.1, 0.2, 0.3, 0.4, and 1) samples with different Fe contents was prepared and investigated as high-capacity cathodic hosts of Na-ion batteries. The partial substitution of Ni and Mn with Fe in the O3-phase lattice can greatly improve the electrochemical performance and the structural stability. A NaFe0.2Mn0.4Ni0.4O2 cathode with an optimized Fe content of x = 0.2 can deliver an initial reversible capacity of 131 mAh g(-1), a reversible capacity greater than 95% over 30 cycles, and a high rate capacity of 86 mAh g(-1) at 10 C in a voltage range of 2.0-4.0 V. The structural characterizations reveal that pristine NaMn0.5Ni0.5O2 and Fe-substituted NaFe0.2Mn0.4Ni0.4O2 lattices underwent different phase transformations from P3 to P3″ and from P3 to OP2 phases, respectively, at high voltage interval. The as-resulted OP2 phase by Fe substitution has smaller interslab distance (5.13 Å) than the P3″ phase (5.72 Å), which suppresses the co-insertion of the solvent molecules, the electrolyte anions, or both and therefore enhances the cycling stability in the high voltage charge. This finding suggests a new strategy for creating cycle-stable transition-metal oxide cathodes for high-performance Na-ion batteries.

National Health and Nutrition Examination Survey whole-body dual-energy X-ray absorptiometry reference data for GE Lunar systems.

The National Health and Nutrition Examination Survey (NHANES 1999-2004) includes adult and pediatric comparisons for total body bone and body composition results. Because dual-energy x-ray absorptiometry (DXA) measurements from different manufacturers are not standardized, NHANES reference values currently are applicable only to a single make and model of Hologic DXA system. The purpose of this study was to derive body composition reference curves for GE Healthcare Lunar DXA systems. Published values from the NHANES 1999-2004 survey were acquired from the Centers for Disease Control and Prevention website. Using previously reported cross-calibration equations between Hologic and GE-Lunar, we converted the total body and regional bone and soft-tissue measurements from NHANES 1999-2004 to GE-Lunar values. The LMS (LmsChartMaker Pro Version 3.5) curve fitting method was used to generate GE-Lunar reference curves. Separate curves were generated for each sex and ethnicity. The reference curves were also divided into pediatric (≤20 years old) and adult (>20 years old) groups. Adult reference curves were derived as a function of age. Additional relationships of pediatric DXA values were derived as a function of height, lean mass, and bone area. Robustness was tested between Hologic and GE-Lunar Z-score values. The NHANES 1999-2004 survey included a sample of 20,672 participants' (9630 female) DXA scans. A total of 8056 participants were younger than 20 yr and were included in the pediatric reference data set. Participants enrolled in the study who weighed more than 136 kg (over scanner table limit) were excluded. The average Z-scores comparing the new GE-Lunar reference curves are close to zero, and the standard deviation of the Z-scores are close to one for all variables. As expected, all measurements on the GE-Lunar reference curves for participants younger than 20 yr increase monotonically with age. In the adult population, most of the curves are constant at younger age and drop moderately as age increases. We have presented NHANES reference curves applicable to DXA whole-body scans acquired on GE Healthcare Lunar systems by age, sex and ethnicity. Users of GE Healthcare GE-Lunar DXA systems can now benefit from the large body composition reference data set collected in the NHANES 1999-2004 study.

Navigation-guided en bloc resection and defect reconstruction of craniomaxillary bony tumours.

The aim of this study was to evaluate the validity of navigation-guided en bloc tumour resection and defect reconstruction in the treatment of craniomaxillary bony tumours. Three patients with ossifying fibroma and two patients with fibrous dysplasia were enrolled in this study. After preoperative planning and three-dimensional simulation, the osteotomy lines for resection were delineated and the normal anatomic structures for defect reconstruction were ascertained. With the guidance of an Accu-Navi navigation system, an en bloc tumour resection and simultaneous defect rehabilitation were performed. The system provided continuously updated information on the position and movement of surgical instruments in the operating field in relation to the preoperative imaging data set. The system error measured by the computer did not exceed 1mm. The osteotomy lines and reconstruction contour were checked by postoperative computed tomography, and good matching with the preoperative planning was achieved. Patients showed no signs of tumour recurrence or prosthesis infection during follow-up (range 12-35 months). Image-guided navigation makes radical bone tumour resection more reliable by implementing preoperative planning, showing the determined safety margins, preserving vital structures and guiding reconstruction.

The association between circulating oestradiol levels and severity of liver disease in males with hepatitis B virus infection.

BACKGROUND: Males have a strikingly increased risk of advanced liver disease. This study was designed to investigate the association between sex hormone levels and different disease states in hepatitis B virus (HBV)-infected male patients. METHODS: Two hundred and fifty-two male patients, including 30 with acute hepatitis B (AHB), 127 with chronic hepatitis B (CHB) and 95 with chronic severe hepatitis B (CSHB), and 48 healthy controls (HCs) were enroled in this study. All patients were followed for 4 months. The primary end point was in-hospital mortality at the end of 90 days. At study entry, a blood sample was collected from all subjects to examine liver function, renal function and sex hormone levels (serum total testosterone and oestradiol). A total of nine clinical chemistry and biochemical variables were analyzed for possible association with outcomes by using Cox proportional hazards and multiple regression models. RESULTS: We demonstrated that in CSHB and CHB patients, oestradiol levels were significantly increased compared with HCs and AHB patients, whereas no significant differences were observed in AHB patients and HCs. All male patients had normal levels of testosterone which were not significantly different compared with those of HCs. Increased oestradiol levels were clinically associated with severe liver disease, and increased 3-month mortality rate. Multivariate analysis demonstrated that oestradiol and the model for end-stage liver disease (MELD) score were independent predictors for mortality (both P < 0.001). CONCLUSION: Oestradiol levels are significantly increased in male patients with hepatitis B, and were associated with the severity of liver disease. Moreover, elevation of oestradiol is an independent predictive factor for the 3-month mortality rate in male patients with hepatitis B.

Study on mechanism of wet air oxidation of emulsification wastewater.

Wet air oxidation (WAO) can effectively be used to treat high-concentration, non-biodegradable emulsification wastewater that contains nonionic matters. Gas chromatograph analysis of emulsification wastewater after oxidation indicated that a catalyst increased production of fatty acids but could not promote its oxidation between 160 and 180 degrees C. When the temperature was greater than or equal to 220 degrees C, the catalyst not only increased production of fatty acids initially but effectively promoted its oxidation in later stages and significantly reduced the concentration of residual surfactants. Experiments proved that fatty acids (especially acetic acid) were the primary intermediate products and that oxidation of these acids was the rate-limiting step. During the process of catalytic WAO of emulsification wastewater, active oxygen molecules attacked organic matters resulting in production of fatty acids, ketone, alcohol, hydrocarbon, and oligo-polyether through radical chain reactions.

Decrease and dysfunction of endothelial progenitor cells in umbilical cord blood with maternal pre-eclampsia.

BACKGROUND: The pre-eclampsia is characterized by placental defective angiogenesis and maternal vascular/endothelial dysfunction. Recently, the decrease and senescence of endothelial progenitor cells (EPC) has been observed in maternal circulation with pre-eclampsia. Given the essential involvement of EPC in neovascularization and reendothelialization, we investigate whether or not the depletion of EPC is existent in placental/fetal circulation with maternal pre-eclampsia. METHODS: Samples of venous cord blood were collected during the labor of preeclamptic mothers (n = 14) and normotensive controls (n = 10). Circulating EPC were enumerated as AC133+/KDR+ cells via fluorescence-activated cell sorting (FACS) analysis. Additionally, EPC were expanded in vitro and identified by DiI-acLDL uptake and lectin staining by direct fluorescent staining under a laser scanning confocal microscope. EPC proliferation, migration and vasculogenesis activities were determined by MTT, modified Boyden chamber assay and in vitro vasculogenensis assay. RESULT: The placental/fetal circulating EPC numbers were significantly decreased in the pre-eclampsia group compared with the control (median, 200; range, 100-440 cells/mL vs 390; 270-440 cells/mL, P < 0.001), and after in vitro cultivation the numbers of EPC also decreased in pre-eclampsia group (19.5; 5.0-32.0 vs 39.5; 31.2-52.0 EPC/x200 field; P < 0.001). Both circulating EPC and cultivated EPC were inversely correlated with cord blood level of soluble fms-like tyrosine kinase 1 (sFlt-1). In addition, the EPC from patients with pre-eclampsia were significantly impaired in their proliferation, migration and vasculogenesis capacities. CONCLUSION: The present study documented the decrease and dysfunction of placental/fetal circulating EPC in patients with pre-eclampsia. The alteration is probably associated with the increased sFlt-1 levels in the umbilical cord blood.

Inhibition of papilloma progression by antisense oligonucleotides targeted to HPV11 E6/E7 RNA.

Human papillomaviruses (HPVs) are recognized as important human pathogens, causing a spectrum of hyperproliferative lesions from benign warts to cervical dysplasias/carcinomas. HPV-associated lesions require continued production of the oncogenic E6/E7 proteins, which are encoded by either bicistronic or overlapping mRNAs. Here we targeted the E6/E7 mRNA of HPV11, a type implicated in causation of genital warts, using molecular reagents. Accessible sites in the HPV11(E6/E7) RNA were identified using library selection protocols, and nucleic acids (DNAzymes, antisense oligonucleotides) targeted to these sites were constructed, and tested in cell culture and on human foreskin grafts. While DNAzymes were at least equally effective in cell culture, antisense oligonucleotides targeted to the region surrounding one of the library-selected sites (ASO(407)) proved most effective in blocking progression of HPV11-induced papillomas in human foreskin grafts on immunodeficient mice. In total, 11 papillomas were treated with ASO(407). Of these, four of seven small papillomas treated with ASO(407) showed loss of detectable virus by in situ hybridization (ISH), and in all four of these, papillomas were no longer evident grossly or histologically after treatment. When larger papillomas were treated, one of four showed loss of virus by ISH, associated with a minor decrease in papilloma size. Considering all 11 papillomas treated with ASO(407), loss of viral staining by ISH was significantly different from that observed in controls (P<0.016), as was true for the seven small treated papillomas (P<0.012). DNAzymes targeted to the same site (or other library selected sites) did not produce statistically significant differences in ISH staining (P<0.15). Our results with ASO(407) appear to represent the first specific molecular therapy against a bona fide HPV infection, and provide a rational proof-of-principle strategy for development of molecular therapeutics targeting other HPV-associated lesions.

Characterization of the rabbit homolog of human MUC1 glycoprotein isolated from bladder by affinity chromatography on immobilized jacalin.

The urinary bladder is lined by transitional epithelium, the glycocalyx on the luminal surface has interesting properties and is implicated in protective functions. Glycoconjugates are major components of the glycocalyx, but their biochemical nature is not well understood. Previous studies on rabbit bladder indicated the presence of significant levels of sialoglycoproteins compared to glycosaminoglycans in the epithelium. In this study, rabbit explant cultures were radiolabeled by precursor sugars or amino acids and a major lectin-reactive glycoprotein of rabbit bladder mucosa was isolated by affinity chromatography on jacalin-agarose. The radiolabeled glycoprotein was purified to homogeneity by a second cycle on the lectin column, followed by gel filtration and density gradient centrifugation. The average molecular mass of the glycoprotein was estimated to be 245 kDa and 210 kDa by gel filtration and SDS-PAGE, respectively. Its buoyant density was 1.40 g/ml, suggesting a carbohydrate content of approximately 50%. The percent distribution of glucosamine-derived tritium label in sialic acid, galactosamine, and glucosamine was 30, 52, and 18, respectively. The glycoprotein consisted entirely of small sialylated and neutral oligosaccharides O-glycosidically linked to serine and threonine residues. The same glycoprotein could be immunoprecipitated with an antibody against the carboxy terminal 17 amino acid peptide of human MUC1 mucin glycoprotein. This suggests that this mucin glycoprotein is the rabbit homolog of MUC1 glycoprotein, which has been previously established to be a component of human bladder urothelium and has been purified from human urine and biochemically characterized.

Lectin histochemical examination of rabbit bladder glycoproteins and characterization of a mucin isolated from the bladder mucosa.

The glycocalyx of the mucosal surface of urinary bladder acts as an effective barrier against invasion by pathogenic microorganisms and injury from toxic substances in the urine. Defects in these bladder mucosal components could thus be important factors in the development of diseases such as interstitial cystitis and lower urinary tract infections. However, information on the nature of glycoconjugates of mammalian bladder mucosa is very limited. In this study, the glycoconjugates of rabbit bladder were examined histochemically using biotinylated lectins with specificities for a variety of carbohydrate moieties. Three [Artocarpus integrifolia (Jacalin), Datura stramonium (DSL), and Maackia amurensis II (MAL-II)] of the lectins bound predominantly to the luminal cell layer, with decreased binding to the basal layers of the epithelium. In contrast, Ricinus communis I and Sambucus nigra lectins did not bind to the cells in the epithelium but strongly interacted with the subepithelial layers, especially the lamina propria. The intensity of the staining by Jacalin and MAL-II was significantly reduced by prior treatment of the bladder sections with O-sialoglycoprotein endopeptidase, indicating that the ligands of these lectins are primarily mucin glycoproteins. In parallel biochemical studies, a high-molecular-weight glycoprotein with characteristics typical of epithelial mucins was purified from the mucosa of rabbit bladder explant cultures metabolically labeled with [(3)H]glucosamine. Quantitative analysis of the sialic acid, uronic acid, and hexosamine contents of delipidated rabbit bladder mucosa revealed a larger proportion of sialoglycoproteins compared with glycosaminoglycans. Taken together, the results of histochemical and biochemical analyses indicate that glycoproteins rather than glycosaminoglycans are the major components of the bladder epithelium, and that the former include a mucin.

Vectors allowing amplified expression of the Saccharomyces cerevisiae Gal3p-Gal80p-Gal4p transcription switch: applications to galactose-regulated high-level production of proteins.

The Gal4, Gal80, and Gal3 proteins of Saccharomyces cerevisiae constitute a galactose-responsive regulatory switch for GAL gene promoters. The low cellular levels of these proteins have hampered mechanistic studies and limit the utility of the GAL gene promoters for high-yield production of endogenous and exogenous proteins. We have constructed two new vectors, pMEGA2 and pMEGA2-DeltaURA3, that increase the level of the Gal4p-Gal80p-Gal3p switch proteins under conditions that preserve the Gal3p-Gal80p-Gal4p stoichiometries required for normal switch function. Cells carrying pMEGA2 show 15- to 20-fold more Gal4p and 30- to 40-fold more Gal3p and Gal80p than cells lacking pMEGA2. These high levels of Gal4p, Gal80p, and Gal3p do not perturb the integrity of galactose-inducible regulation. Cells that carry pMEGA2 exhibit normal galactose-induction kinetics for the chromosomal MEL1 gene expression and normal, albeit slower, log-phase growth. Insertion of the MEL1 gene into pMEGA2 provides a 24- to 30-fold increase in the Mel1 protein. Cells carrying a 2-microm-based URA3-selectable plasmid containing a GAL1pro:lacZ reporter gene and a second plasmid, pMEGA2-DeltaURA3, produce 12-fold more beta-galactosidase than cells carrying only the GAL1pro:lacZ reporter plasmid. The performance of the MEGA plasmids in providing amplified production of the Gal3, Gal80, and Gal4 proteins should prove useful in investigations of the mechanistic aspects of these transcription switch proteins and in work aimed at achieving high-level, galactose-regulatable production of proteins in yeast.