Relationships between Single Nucleotide Polymorphism Markers and Meat Quality Traits of Duroc Breeding Stocks in Korea.

This study was conducted to determine the relationships of five intragenic single nucleotide polymorphism (SNP) markers (protein kinase adenosine monophosphate-activated γ3 subunit [PRKAG3], fatty acid synthase [FASN], calpastatin [CAST], high mobility group AT-hook 1 [HMGA1], and melanocortin-4 receptor [MC4R]) and meat quality traits of Duroc breeding stocks in Korea. A total of 200 purebred Duroc gilts from 8 sires and 40 dams at 4 pig breeding farms from 2010 to 2011 reaching market weight (110 kg) were slaughtered and their carcasses were chilled overnight. Longissimus dorsi muscles were removed from the carcass after 24 h of slaughter and used to determine pork properties including carcass weight, backfat thickness, moisture, intramuscular fat, pH24h, shear force, redness, texture, and fatty acid composition. The PRKAG3, FASN, CAST, and MC4R gene SNPs were significantly associated with the meat quality traits (p<0.003). The meats of PRKAG3 (A 0.024/G 0.976) AA genotype had higher pH, redness and texture than those from PRKAG3 GG genotype. Meats of FASN (C 0.301/A 0.699) AA genotype had higher backfat thickness, texture, stearic acid, oleic acid and polyunsaturated fatty acid than FASN CC genotype. While the carcasses of CAST (A 0.373/G 0.627) AA genotype had thicker backfat, and lower shear force, palmitoleic acid and oleic acid content, they had higher stearic acid content than those from the CAST GG genotype. The MC4R (G 0.208/A 0.792) AA genotype were involved in increasing backfat thickness, carcass weight, moisture and saturated fatty acid content, and decreasing unsaturated fatty acid content in Duroc meat. These results indicated that the five SNP markers tested can be a help to select Duroc breed to improve carcass and meat quality properties in crossbred pigs.

Antioxidant Enzyme Activity and Meat Quality of Meat Type Ducks Fed with Dried Oregano (Origanum vulgare L.) Powder.

One-day-old Cherry valley meat-strain ducks were used to investigate the effect of supplemental dried oregano powder (DOP) in feed on the productivity, antioxidant enzyme activity, and breast meat quality. One hundred sixty five ducks were assigned to 5 dietary treatments for 42 days. The dietary treatment groups were control group (CON; no antibiotic, no DOP), antibiotic group (ANT; CON+0.1% Patrol), 0.1% DOP (CON+0.1% DOP), 0.5% DOP (CON+0.5% DOP), and 1.0% DOP (CON+1.0% DOP). Upon feeding, 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radical scavenging activity of oregano extracts was higher than that of tocopherol, although it was less than that of ascorbic acid. As a result of in vivo study, DOP in the diet showed no effects on final body weight, feed intake, or feed conversion ratio. However, dietary 0.5% and 1% DOP supplementation caused a significant increase in the serum enzyme activity of superoxide dismutase (SOD) compared with CON and ANT, while glutathione peroxidase (GPx) in tissue was increased as compared to ANT (p<0.05). Cooking loss from ducks fed with DOP decreased compared with the control ducks. Thiobarbituric acid reactive substance (TBARS) values of duck breast meat at 5 d post slaughter was found to be significantly reduced in ducks whose diets were supplemented with 0.5% and 1% DOP (p<0.05). These results suggest that diets containing 0.5% and 1% DOP may beneficially affect antioxidant enzyme activity of GPx and SOD, improve meat cooking loss, and reduce TBARS values in breast meat at 5 d of storage in ducks.

Effects of oxygenated or hydrogenated water on growth performance, blood parameters, and antioxidant enzyme activity of broiler chickens.

This study was conducted to investigate the effects of providing oxygenated and hydrogenated water on the growth performance, blood biochemical parameters, and immunoglobulin concentrations and antioxidant enzyme activity of broiler chickens. In our investigation, 144 Ross × Ross broiler chicks were randomly allotted to three different treatment groups with four replicates (treatment × replicate × bird = 3 × 4 × 12). All chicks were given one of the following types of water for five weeks: tap water (CON), hydrogenated water (HNW), and oxygenated water (ONW). ONW supplementation increased the final body weight and weight gain and also improved both feed intake and feed conversion of broiler chickens as compared to those of CON broiler chickens (P < 0.05). The abdominal fat and its ratio to the final body weight showed that fat accumulation in the broiler chicken abdomen was reduced when broiler chickens drank only ONW for five weeks (P < 0.05). ONW supplementation improved blood parameters, including triacylglyceride, total cholesterol, and low-density lipoprotein-cholesterol. Additionally, in accordance with a globulin increase in broiler chickens, both IgG and IgM generation were significantly enhanced when ONW was supplied to broiler chickens (P < 0.05) but only a numerical advance was observed in the HNW group (P > 0.05). Both oxygenated and hydrogenated water supplementation significantly improved the antioxidant effects (P < 0.05), and it seems that superoxide dismutase refinement was completed due to oxygen and/or hydrogen enhancement of drinking water. These results indicate that oxygen enhancement of drinking water may be recommended to improve growth performance by increasing immunoglobulins mainly IgG and IgM.

Cyclic fatigue resistance, torsional resistance, and metallurgical characteristics of V taper 2 and V taper 2H rotary NiTi files.

The aim of this study was to compare the cyclic fatigue resistance, torsional resistance, and metallurgical characteristics of conventional NiTi wire (V taper 2, V2) and CM wire (V taper 2H, V2H)-based files. Cyclic fatigue and torsional resistance of V2 and V2H were investigated by measuring the number of cycles to fracture, maximum torque at fracture, and maximum angle at fracture. The typical patterns of fatigue and torsional fractures were investigated using a scanning electron microscope (SEM). The metallurgical characteristics were investigated by differential scanning calorimetry (DSC) from -100 °C to 100 °C. The austenite finishing temperature (Af) of each instrument was also measured. The microstructures of the instruments were investigated by a transmission electron microscope (TEM) along with selected area diffraction pattern analysis. The results were statistically analyzed by Mann-Whitney U-test (p = 0.05). V2H showed significantly higher cyclic fatigue resistance and torsional resistance than V2. SEM images of the fractured surfaces showed typical patterns of fatigue and torsional fracture. The DSC analysis of V2 showed one small peak in both the heating and cooling curves. The Af of V2 was -0.32 °C. V2H showed two remarkable peaks in the heating curve and one remarkable peak in the cooling curve. The Af of V2H was 33.25 °C. The TEM analysis showed that both V2 and V2H are mainly composed of austenite. In conclusion, V2H showed higher cyclic fatigue resistance and torsional resistance than V2. The superior properties of V2H could be attributed to the annealing effect and possibly the martensite phase. SCANNING 38:564-570, 2016. © 2016 Wiley Periodicals, Inc.

Aberrant protein expression of transcription factors BACH1 and ERG, both encoded on chromosome 21, in brains of patients with Down syndrome and Alzheimer's disease.

Down syndrome (DS; trisomy 21) is a genetic disorder associated with early mental retardation and patients inevitably develop Alzheimer's disease (AD)-like neuropathological changes. The molecular defects underlying the DS-phenotype may be due to overexpression of genes encoded on chromosome 21. This so-called gene dosage hypothesis is still controversial and demands systematic work on protein expression. A series of transcription factors (TF) are encoded on chromosome 21 and are considered to play a pathogenetic role in DS. We therefore decided to study brain expression of TF encoded on chromosome 21 in patients with DS and AD compared to controls: Frontal cortex of 6 male DS patients, 6 male patients with AD and 6 male controls were used for the experiments. Immunoblotting was used to determine protein levels of TF BACH1, ERG, SIM2 and RUNX1. SIM2 and RUNX1 were comparable between groups, while BACH1 was significantly reduced in DS, and ERG was increased in DS and AD as compared to controls. These findings may indicate that DS pathogenesis cannot be simply explained by the gene dosage effect hypothesis and that results of ERG expression in DS were paralleling those in AD probably reflecting a common pathogenetic mechanism possibly explaining why all DS patients develop AD like neuropathology from the fourth decade. We conclude that TF derangement is not only due to the process of neurodegeneration and propose that TFs BACH1 and ERG play a role for the development of AD-like neuropathology in DS and pathogenesis of AD per se and the manifold increase of ERG in both disorders may form a pivotal pathogenetic link.

Reduction of chromatin assembly factor 1 p60 and C21orf2 protein, encoded on chromosome 21, in Down syndrome brain.

Trisomy 21 (Down syndrome, DS) is the most common genetic cause of mental retardation, resulting from triplication of the whole or distal part of human chromosome 21. Overexpression of genes located on chromosome 21, as a result of extra gene load, has been considered a central hypothesis for the explanation of the DS phenotype. This gene dosage hypothesis has been challenged, however. We have therefore decided to study proteins whose genes are encoded on chromosome 21 in brain of patients with DS and Alzheimer's disease (AD), as all patients with DS from the fourth decade show Alzheimer-related neuropathology. Using immunoblotting we determined Coxsackievirus and adenovirus receptor (CAR), Claudin-8, C21orf2, Chromatin assembly factor 1 p60 subunit (CAF-1 p60) in frontal cortex from DS, AD and control patients. Significant reduction of C21orf2 and CAF-1 p60, but comparable expression of CAR and claudin-8 was observed in DS but all proteins were comparable to controls in AD, even when related to NSE levels to rule out neuronal cell loss or actin to normalise versus a housekeeping protein. Reduced CAF-1 p60 may reflect impaired DNA repair most probably due to oxidative stress found as early as in fetal life continuing into adulthood. The decrease of C21orf2 may represent mitochondrial dysfunction that has been reported repeatedly and also data on CAR and claudin-8 are not supporting the gene-dosage hypothesis at the protein level. As aberrant expression of the four proteins was not found in brains of patients with AD, decreased CAF and C21orf2 can be considered specific for DS.

Hepatitis C in liver allografts.

Recurrent or intercurrent hepatitis C represents significant problems in patients with liver transplants and must be differentiated from hepatic allograft rejection and other conditions affecting allografts. Often, the currently available anti-hepatitis C virus (HCV) tests are not helpful in the differential diagnosis, because preexisting anti-HCV may persist after transplantation or its development may be delayed. We determined the presence of HCV RNA by the reverse double polymerase chain reaction in biopsy specimens of liver allografts from nine patients with acute or chronic hepatitis of uncertain origin and from three patients with cellular allograft rejection. The NS3 region sequences of HCV were detected in seven of nine liver allograft biopsy specimens 6 weeks to 20 months after transplantation. Hepatitis C virus RNA was not detected in two patients. One of these patients was anti-HCV positive, showing mild acute hepatitis 5 weeks after transplantation. Anti-HCV was present in three patients with detectable HCV RNA in the liver but was absent from four other patients with HCV RNA. These findings suggest that HCV is a major cause of acute and chronic hepatitis following liver transplantation, that detection of HCV RNA by polymerase chain reaction in the liver biopsy specimen represents a reliable method for the diagnosis of hepatitis C in liver allografts, and that in some patients HCV may be acquired during transplantation while in others it may represent a recurrent infection.

The effects of long-term administration of oral desmopressin on the baseline secretion of antidiuretic hormone and serum sodium concentration for the treatment of nocturia: a circadian study.

PURPOSE: We assessed the effects of long-term oral desmopressin on serum sodium and baseline antidiuretic hormone secretion in elderly patients with nocturia. MATERIALS AND METHODS: A total of 15 elderly male patients with severe nocturia (greater than 3 voids nightly) who did not show hyponatremia within 7 days of administration of 0.2 mg desmopressin were enrolled in this study. Desmopressin (0.2 mg) was administered orally nightly for 1 year. Before and 1 month after the 1-year medication 24-hour circadian studies were performed to monitor changes in antidiuretic hormone. Every 3 months during the 1-year medication serum changes and timed urine chemistry were monitored. RESULTS: Desmopressin significantly decreased nocturnal urine output and the number of nocturia episodes (p<0.01). Compared to before treatment desmopressin gradually decreased serum sodium and induced statistically but not clinically significant hyponatremia after 6 months of treatment. After discontinuing desmopressin serum sodium returned to the normal range in all patients. There were no significant differences when baseline and posttreatment endogenous antidiuretic hormone were compared. No serious systemic complications were found during medication. CONCLUSIONS: Long-term desmopressin administration gradually decreased the serum concentration and induced significant hyponatremia from 6 months in patients who did not show initial hyponatremia. Long-term administration of desmopressin for 1 year in elderly patients did not affect baseline antidiuretic hormone secretion. For long-term desmopressin administration serum sodium should be assessed regularly, at least every 6 months.

Analysis of differentially expressed transcripts from planthopper-infested wild rice (Oryza minuta).

A subtracted library was constructed from planthopper-infested wild rice (Oryza minuta) by suppression subtractive hybridization in combination with mirror orientation selection. To screen the differentially expressed transcripts in the library, we applied a cDNA microarray containing 960 random clones in a reverse Northern blot analysis using cDNA probes prepared from the mRNAs of control and planthopper-infested samples. On the basis of the signal intensities and expression ratios obtained from experiments performed in triplicate, we selected 383 clones. The elevated expression levels and overall profiles over time were verified by Northern blot analysis. Although Southern blot analysis showed similar copy numbers of the screened genes in O. minuta and O. sativa, it also revealed that the expression profiles had a different pattern. Functional categorization placed the identified transcripts in the categories of subcellular localization, metabolism, and protein fate. The presence of these expressed sequence tags implies that resistance of O. minuta to insect infestation can be achieved not only by an elevated expression of defense-related genes but also by enhanced metabolic activities.

Comparative studies on the detection of hepatitis B virus DNA in frozen and paraffin sections by the polymerase chain reaction.

The polymerase chain reaction (PCR) is an extremely sensitive technique that has been used for detection of DNA sequences in formalin-fixed, paraffin-embedded tissues. In order to verify that hepatitis B virus (HBV) DNA sequences are adequately preserved in routinely processed liver tissues, we performed PCR with five different primer pairs for HBV sequences on DNA extracted by two different methods from paraffin and frozen liver sections. The amount of PCR products obtained with DNA templates extracted by the proteinase K-SDS method from frozen sections was significantly larger than that from paraffin sections. However, boiling of deparaffinized sections in water containing Chelex-100 resulted in ample amounts of PCR products irrespective of the primers used. On Southern blots, the location of the bands of amplified DNA obtained by the different methods was consistent with the predicted size, suggesting that the viral sequences had not been altered by processing. Although freezing of fresh tissue yields quantitatively more HBV DNA, formalin fixation qualitatively preserves the viral DNA sequences adequately for detection by PCR. Therefore, formalin-fixed, paraffin-embedded tissues may be used for the detection of viral DNA sequences by PCR. Application of the described procedure to routinely processed tissues significantly broadens the applicability of this powerful diagnostic and investigative method.

The interaction of DNA mismatch repair proteins with human exonuclease I.

Exonucleolytic degradation of DNA is an essential part of many DNA metabolic processes including DNA mismatch repair (MMR) and recombination. Human exonuclease I (hExoI) is a member of a family of conserved 5' --> 3' exonucleases, which are implicated in these processes by genetic studies. Here, we demonstrate that hExoI binds strongly to hMLH1, and we describe interaction regions between hExoI and the MMR proteins hMSH2, hMSH3, and hMLH1. In addition, hExoI forms an immunoprecipitable complex with hMLH1/hPMS2 in vivo. The study of interaction regions suggests a biochemical mechanism of the involvement of hExoI as a downstream effector in MMR and/or DNA recombination.

Protein levels of genes encoded on chromosome 21 in fetal Down syndrome brain: Challenging the gene dosage effect hypothesis (Part IV).

Down syndrome (DS) is the most frequent genetic disorder with mental retardation and caused by trisomy 21. Although the molecular mechanisms of the various phenotypes of DS could be due to overexpression of gene(s) on chromosome 21, several groups have challenged this gene dosage effect hypothesis. The near completion of the sequencing of human chromosome 21 provides unprecedented opportunities to understand the molecular pathology of DS, however, functional information on gene products is limited so far. We therefore evaluated the levels of six proteins whose genes are encoded on chromosome 21 (trefoil factor 1, trefoil factor 2, trefoil factor 3, coxsackie virus and adenovirus receptor, carbonyl reductase 1 and interferon- alpha receptor) in fetal cerebral cortex from DS and controls at the early second trimester using Western blot analysis. None of the investigated proteins showed overexpression in DS compared to controls suggesting that these proteins are not involved in abnormal development of fetal DS brain and that DS phenotype can not be simply explained by the gene dosage effect hypothesis. We are systematically quantifying all proteins whose genes are encoded on chromosome 21 and these studies may provide a better understanding of genotype-phenotype correlation in DS.

Elevated serum levels of transforming growth factor-beta1 in patients with colorectal carcinoma: its association with tumor progression and its significant decrease after curative surgical resection.

BACKGROUND: Transforming growth factor-beta1 (TGF-beta1) acts as a potent inhibitor of cell growth and tumor progression but loss of this negative regulation can contribute to tumor development. Some studies have reported an association between disease progression and TGF-beta1 expression in patients with colorectal carcinoma, but their results were not always consistent. METHODS: Serum levels of TGF-beta1 were measured using an enzyme-linked immunoadsorbent assay in 121 consecutive patients with colorectal carcinoma and compared with TGF-beta1 serum levels in 31 healthy volunteers. Serum levels of TGF-beta1 also were measured in 50 patients who underwent curative surgical resection (part of the 121 preoperative patients) to compare their levels with preoperative serum levels of TGF-beta1. RESULTS: Serum levels of TGF-beta1 in patients with colorectal carcinoma (45+/-15 ng/mL) (mean+/-the standard deviation) were significantly higher than those in the healthy control group (32+/-4 ng/mL) (P = 0.001). Serum levels of TGF-beta1 increased with increasing tumor stage (P < 0.01). Serum levels of TGF-beta1 were correlated significantly with depth of tumor invasion, lymph node metastasis, distant metastasis, and serum levels of carcinoembryonic antigen (CEA). Serum levels of TGF-beta1 tended to increase with increasing CEA (correlation coefficient = 0.21; P < 0.05). The mean serum level of TGF-beta1 in patients with colorectal carcinoma before surgery (45+/-14 ng/mL) (n = 50) significantly decreased to 34+/-7 ng/mL, which was within the normal range (32+/-4 ng/mL), after curative surgical resection of the tumor (P = 0.0000). Serum levels of TGF-beta1 after tumor resection decreased more significantly in patients with higher preoperative levels of TGF-beta1 (from 53+/-12 ng/mL to 36+/-6 ng/mL) (n = 30). CONCLUSIONS: The results of the current study suggest that serum levels of TGF-beta1 in colorectal carcinoma patients may be associated with disease progression and may be used as a biomarker in the management of colorectal carcinoma patients. The authors believe further studies with a large number of patients for a longer follow-up period are necessary to conclude whether serum levels of TGF-beta1 carry significant clinical relevance.

Effect of short-term prednisolone therapy in patients with severe chronic type B hepatitis.

Ten patients with severe chronic type B hepatitis confirmed by liver biopsy were treated with prednisolone for eight weeks and followed up for more than one year. The patients were comprised of 6 males and 4 females, ages 17 to 45 (mean 32) yrs. Serum alanine aminotransferase (ALT) was elevated more than one month before the treatment in all (mean: 379 U/L, range: 87 to 772 U/L). Initial serological tests showed hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in all and hepatitis B virus DNA (HBV-DNA) in 7/10 (70%). Liver biopsy showed severe chronic active hepatitis with confluent necrosis or acinar hepatitis in all. Prednisolone, 60 mg/day, was administered initially and the dose was tapered every 2 weeks over the 8 weeks period. Two to six months after cessation of treatment, 5 of 10 patients showed a disappearance of HBeAg and serum HBV-DNA and return of serum ALT level to normal (responders). The initial serum ALT level in responders was slightly higher than that of non-responders (mean: 404 vs. 355 U/L), but there was no statistical significance. Among 5 responders, serum HBV-DNA was detected in three patients initially and was transiently detected in one patient during treatment. In non-responders, HBeAg persisted during and after the treatment and serum HBV-DNA persisted in three, but serum ALT was decreased in all. One patient who did not show any clinical or serological improvement, died of jaundice, ascites and hepatic encephalopathy 4 months later.(ABSTRACT TRUNCATED AT 250 WORDS)

Three-dimensional demonstration and endoscopic treatment of pancreaticoperitoneal fistula.

An alcoholic man presented with bloody ascites, which was confirmed as pancreatic ascites complicating chronic pancreatitis. Endoscopic retrograde pancreatography [ERP] demonstrated a ductal disruption at the head of the pancreas, a fistulous tract, and extravasation to the peritoneal cavity. Furthermore, a computerized tomographic scan subsequent to the endoscopic retrograde pancreatography (ERP-CT scan) gave the three-dimensional anatomy of the fistulous tract by the residual contrast media in the pancreatic duct and the fistulous tract. The pancreatic ascites, which was refractory to conventional medical treatment of a 5-wk duration, was successfully treated by endoscopic placement of a pancreatic stent and administration of a somatostatin analogue.

Detection of replicative hepatitis C virus sequences in hepatocellular carcinoma.

Several serologic studies suggest that infection by hepatitis C virus (HCV) may be associated with the development of hepatocellular carcinoma (HCC). Therefore, we examined tumor tissue and/or the surrounding liver of 20 patients for viral sequences by the polymerase chain reaction (PCR). In 12 cases, liver and tumor tissues were separable for extraction. RNA was extracted from frozen tissues and used as a template for reverse transcription followed by double PCR with nested primers for the 5'-untranslated (NT) and nonstructural NS3 regions of HCV. In addition, the tissue extracts were tested by single PCR for X gene and S gene sequences of hepatitis B virus (HBV). NT region sequences of HCV were detected in the available tumor tissue of all anti-HCV-positive patients except for one. Negative (replicative) strands of HCV RNA were found in the same tissues as positive (genomic) strands at almost the same relative amounts, suggesting replication of HCV in the tumor tissue rather than contamination by HCV-positive blood. HBV X and S sequences were demonstrated in two tumors, but were absent from three tumors that were surrounded by liver tissues with HBV X sequences. One patient had nucleic acids of both viruses in tumor tissue. These observations suggest that in addition to HBV, HCV may play a role in the development of hepatocellular carcinoma.

Hepatitis C RNA in liver of chronic hepatitis C patients before and after interferon alfa treatment.

BACKGROUND: Successful treatment of chronic hepatitis C with interferon alfa is frequently followed by relapse. Because loss of hepatitis C viral RNA (HCV-RNA) in serum is not predictive of sustained response, the loss of HCV-RNA in liver as a predictor of sustained response was investigated. METHODS: Twenty-one patients with chronic hepatitis C treated with recombinant interferon alpha had HCV-RNA sequences determined in frozen liver tissue before and after treatment and in serum at the end of treatment. Reverse double polymerase chain reaction was used to detect sequences to the 5' nontranslated region of the HCV genome using double nested primers. RESULTS: HCV-RNA disappeared in the liver in 10 of 11 (91%) complete responders whereas it remained detectable in the liver or serum of 7 of 8 (87%) nonresponders. Five complete responders relapsed biochemically during 6 month's follow-up; 4 of these had no detectable HCV-RNA in liver at end of treatment. CONCLUSIONS: Disappearance of HCV-RNA in liver correlates with initial clinical outcome, but as previously reported with serum HCV-RNA, this loss does not necessarily allow prediction of a sustained response.

The discomallear ligament and the anterior ligament of malleus: an anatomic study in human adults and fetuses.

According to some reports, movement of the malleus, resulting from anterior hypertension on the discomallear ligament (DML), could produce aural symptoms related with damage to middle ear structures. The aim of this study was to examine the topographic relationship of the DML and the anterior ligament of malleus (ALM). Four fetuses and 16 adult hemi-sectioned heads were used to determine the anatomic-clinical relevance of DML and ALM in temporomandibular disorder. In fetal specimens, the DML was distinctly interposed between the malleus and the disc of the temporomandibular joint (TMJ), and the ALM had a structure apparently composed of the superior and inferior lamellae, running anteriorly in continuation with the sphenomandibular ligament (SML) through the future petrotympanic fissure (PTF). In all adult specimens, the DML was inserted into the malleus, and it expanded broadly toward the disc and capsular region of the TMJ in a triangular shape and inserted into the disc and capsule of the TMJ. The two-lamellae structure of the ALM was not distinguishable in adult specimens. The overstretched ALM resulted in movement of the malleus in five cases, but similar tension applied to the DML did not cause any movement of the malleus. This result provides an indication of the clinical significance of the ALM, a ligamentous structure continuous with the SML. It is apparent that the ALM has the potential to cause aural symptoms as a result of damage to the middle ear structure.

Structure and properties of the 26S protease complex from chick skeletal muscle.

The 26S protease complex was purified from chick skeletal muscle and shown to consist of unusually heterogeneous 21-140 kDa polypeptides, including the 21-32 kDa subunits of the 20S proteasome. Electron microscopic analysis revealed that the 26S complex may have a symmetric morphology with two large rectangular terminal domains attached to a thinner central 20S proteasome domain. The 26S complex was capable of degrading the peptide substrates of the 20S proteasome, including Suc-LLVY-AMC, N-Cbz-LLE-NA and N-Cbz-ARR-MNA. The two enzyme complexes showed similar sensitivities to various site-specific protease inhibitors, although their sensitivities to SDS were differed from each other. Immunoprecipitation with anti-26S complex antibody reduced peptide hydrolysis by the 20S proteasome. Similarly, anti-20S proteasome antibody inhibited peptide hydrolysis by the 26S complex. These results demonstrate that the 26S protease complex contains the 20S proteasome as a functional and structural component.

Increased serum levels of mutant p53 proteins in patients with colorectal cancer.

We have examined the serum levels of the mutant p53 protein in patients with colorectal cancer preoperatively (n=50), and in patients with adenomatous polyp (n=13). Mutant p53 protein in patients after curative surgical resection of colorectal cancer (n=26, part of the fifty preoperative patients) was also measured. Serum samples were stored frozen at -70 degrees C until the time of analysis. We used the p53 mutant ELISA (QIA03, CALBIOCHEM) system. Serum levels of the mutant p53 protein in patients with colorectal cancer (mean=0.97+/-0.14 ng/ml, ranged from 0.7 ng/ml to 1.37 ng/ml, n=50) were significantly greater than those in patients with adenomatous polyp (mean=0.73+/-0.06 ng/ml, ranged from 0.69 ng/ml to 0.83 ng/ml) (p<0.001). There was a significant correlation between serum p53 levels and CA19-9 levels (p<0.01). Serum levels of the mutant p53 protein prior to surgery (mean=0.97+/-0.13 ng/ml, n=26) significantly decreased after surgical resection of tumor (mean=0.82+/-0.07 ng/ml) (p<0.001, paired t-test). These results suggest that mutant p53 protein might be used as a potential biomarker in the management of patients with colorectal cancer. Further study is warranted to establish its clinical significance.