Prediction of Severe Esophageal Varices in Patients With Cirrhosis Based on Levitt's CO Breath Test: A Proof of Concept Study.

GOALS: This study investigated the feasibility of using erythrocyte (RBC) lifespan determined by Levitt's CO breath test (LCOBT) to predict esophageal varices needing treatment (VNT) in patients with cirrhosis. BACKGROUND: Esophageal varix bleeding is a common fatal complication of cirrhosis and portal hypertension. The gold standard for identifying VNT is esophagogastroduodenoscopy (EGD), an invasive procedure with low patient compliance. VNT screening based on Baveno VI criteria has mediocre specificity. STUDY: RBC lifespan was determined by LCOBT in 53 cirrhotic patients (13 without varices, 11 mild/moderate varices, and 29 severe varices). Correlation of varix severity with RBC lifespan and other variables was analyzed. Rates of shortened RBC lifespan and thrombocytopenia (Baveno VI criteria) were compared. RESULTS: RBC lifespan correlated inversely with severity of varices ( r =-0.793, P <0.001). Mean RBC lifespans were 129±31, 96±21, and 59±21 days for Nonvarix, Mild/Moderate, and Severe groups. Shortened RBC lifespan (<75 d) was observed in 79.3% (23/29) of patients with severe varices, a frequency similar or identical to thrombocytopenia rates [original Baveno VI criteria, 86.2% (25/29), P =0.487; expanded criteria, 79.3% (23/29), P >0.999]. Among 24 patients without severe varices, shortened RBC lifespan was observed in 1 patient whereas thrombocytopenia was detected in 13 and 8 patients based on the original ( P <0.001) and expanded criteria ( P =0.010), respectively. CONCLUSIONS: RBC lifespan correlates inversely with varix severity in patients with cirrhosis. LCOBT may enable specific screening for VNT.

Levitt's CO breath test in the differential diagnosis of chronic isolated hyperbilirubinemia.

A key component of the differential diagnosis of isolated hyperbilirubinemia (HB) is distinguishing between hemolytic and non-hemolytic types. Routine hemolysis screening markers have unsatisfactory sensitivity and specificity. Erythrocyte (RBC) lifespan shortening, the gold standard marker of hemolysis, is seldomly measured due to the cumbersome and protracted nature of standard methods. A new Levitt's CO breath test method may enable simple, rapid RBC lifespan measurement. In this pilot prospective diagnostic study, Levitt's CO breath test was evaluated to discriminate hemolytic from non-hemolytic HB in adults. One hundred and thirty eligible non-smoking adult patients who were aged 18 or older, referred for chronic (>6 months) isolated HB or had a known diagnosis of isolated HB of a rare cause, were recruited, including 77 with non-hemolytic HB and 53 with hemolytic HB. ROC curve analysis was applied to determine the optimal cutoff for discriminating between hemolytic and non-hemolytic HB, and the performance was calculated. Results showed that the mean RBC lifespan in non-hemolytic HB (93 ± 26 d) was reduced (p= 0.001 vs. normal reference value of 126 d), but longer than that in hemolytic HB (36 ± 17 d;p= 0.001). RBC lifespans did not differ significantly between 26 patients with simple hemolytic HB (32 ± 14 d) and 27 patients with a Gilbert syndrome comorbidity (40 ± 18 d). ROC curve analysis revealed an optimal lifespan cutoff for discriminating between hemolytic and non-hemolytic HB of 60 d (AUC = 0.982), with a diagnostic accuracy of 95.4%, 94.3% sensitivity and 96.1% specificity respectively. These results indicate that Levitt's CO breath test seems to be very sensitive and specific for detecting hemolysis in adult patients with chronic isolated HB, and could enable simple, rapid, and reliable differential diagnosis of isolated HB. A large-scale validation study of the method is warranted.

Hemodiafiltration improves red blood cell lifespan in patients with end-stage renal disease.

INTRODUCTION: Uremic toxin-induced shortening of red blood cell (RBC) lifespan is an important mechanism of anemia in end-stage renal disease (ESRD). Conventional hemodialysis does not improve RBC lifespan; the efficacy of hemodiafiltration (HDF) for alleviating RBC lifespan has not yet been evaluated in patients with ESRD. METHODS: Twenty-three patients with ESRD in maintenance hemodialysis were enrolled. Baseline data for sex, age, dialysis vintage, pre-dialysis hemoglobin (Hb), blood urea nitrogen (BUN), intact parathyroid hormone (iPTH), single pool Kt/V (spKt/V), and plasma indophenol sulfate (IS) were collected. RBC lifespans before and after one session of HDF were compared. The resultant differences were subjected to correlational analyses with baseline data. RESULTS: RBC lifespan increased from 73 (66, 89) days at baseline to 77 (71, 102) days after a single HDF treatment (p = 0.034). Meanwhile, plasma IS concentration decreased from 113.05 (80.67, 133.05) mg/L to 83.87 (62.98, 96.78) mg/L (p < 0.001). RBC lifespan increases correlated negatively with Hb levels. CONCLUSIONS: A single HDF treatment improved RBC lifespan in ESRD patients on maintenance hemodialysis, with more severe pre-HDF anemia at baseline being associated with greater increases in RBC lifespan.

A Validation Study of a Locally Adapted Brussels Infant and Toddler Stool Scale of the Chinese Version.

The Brussels Infant and Toddler Stool Scale was developed to improve the reliability of constipation diagnosis in non-toilet-trained children. The aim of this study was to evaluate the validity of simplified Chinese versions of the Brussels Infant and Toddler Stool Scale when used by parents, community doctors, pediatricians, and nurses. Photographs of the Scale were categorized into four categories (hard stools, formed stools, loose stools, and watery stools) and subjects assigned each photograph to a category. The study included two stages. In the first stage (n = 237 observers), percent correct allocations of the seven photographs ranged from 68.4% to 93.2%. We observed poorer recognition of the three hard stool items (77.4%, 85.8%, and 74.0%) than had been reported in the original Brussels Infant and Toddler Stool Scale validity study (95.9%, 93.4%, and 96.2%). Because hard stool items were commonly miscategorized as formed stools (21.6%, 9.5%, and 26.0%), we modified the descriptors "hard stools" and "formed stools" into "dry/hard stools" and "formed loose stools," respectively, and examined the performance of the modified Chinese Brussels Infant and Toddler Stool Scale in stage 2 of our study. The proportions of correct allocations of the three "hard stool" items in the modified Chinese Brussels Infant and Toddler Stool Scale increased to 94.7%, 90.4%, and 84.6%, values that were statistically similar to those reported previously in the original Brussels Infant and Toddler Stool Scale publisher. Renaming these categories to remove ambiguity in Chinese improved the identifiability of these items. The resultant Chinese Brussels Infant and Toddler Stool Scale was found to be valid for use with Chinese observers.

Abnormally Short Erythrocyte LifeSpan in Three Patients with Primary Myelofibrosis Despite Successful Control of Splenomegaly.

Splenectomy is an elective operation for refractory anemia in patients with primary myelofibrosis (PMF). We found that 3/3 patients with PMF in our department continued to have very shortened erythrocyte (RBC) lifespans (35 days, 66 days, and 37 days, respectively) after treatment-alleviated splenomegaly. These outcomes suggest that intravascular hemolysis predominantly independent of hypersplenism may underlie, at least to some extent, peripheral hemolysis in patients with PMF. More cases studies are needed to elucidate the role of splenomegaly in PMF-associated anemia.

Gilbert's syndrome coexisting with hereditary spherocytosis might not be rare: Six case reports.

BACKGROUND: Both Gilbert's syndrome (GS) and hereditary spherocytosis (HS) are common genetic disorders. However, comorbidity of GS with HS has always been considered a rare phenomenon, and it can impede accurate diagnoses in the presence of isolated unconjugated hyperbilirubinemia. CASE SUMMARY: In a study on Levitt's carbon monoxide (CO) breath test for the differential diagnosis of isolated hyperbilirubinemia, we found six GS patients with HS in 6 mo. The patients, including five males and one female, aged 25-58 years, were from four families and generally in good health. Their chronic fluctuating jaundice and/or hyperbilirubinemia had been diagnosed as simple constitutional jaundice for 6-30 years. Liver function tests showed isolated unconjugated hyperbilirubinemia with serum total bilirubin ranging from 20.7-75.4 µmol/L. Blood hemoglobin was normal in five cases, and slightly decreased in one (11.5 g/dL). Overt hemolytic signs were absent, while erythrocyte lifespan determined by the newly developed Levitt's CO breath test was significantly short (15-50 d), definitely demonstrating the presence of hemolysis. Given that their unconjugated hyperbilirubinemia compared inappropriately with hemolytic severity, as indicated by the hemoglobin level, further combined genetic tests for both UGT1A1 and hereditary erythrocyte deficiencies were conducted. These tests confirmed, at last, the coexistence of GS with HS. CONCLUSION: Comorbidity of GS and HS might not be uncommon in isolated unconjugated hyperbilirubinemia. While CO breath test would sensitively detect the hemolysis, the discordance between the hyperbilirubinemia and hemoglobin level could strongly indicate the coexistence of GS and HS.

Carbon monoxide breath test assessment of mild hemolysis in Gilbert's syndrome.

BACKGROUND: Mild hemolysis is difficult to determinate by traditional methods, and its role in Gilbert's syndrome (GS) is unclear. The main aims were to inspect the erythrocyte (RBC) survival in GS by using Levitt's carbon monoxide (CO) breath test and to assess its contribution to unconjugated hyperbilirubinemia. METHODS: Fifty subjects with GS and 1 with type-II Crigler-Najjar syndrome (CN2) received RBC lifespan measurement with Levitt's CO breath test. Mean RBC lifespan was compared with normal referral value. Correlations of serum total bilirubin (TB) with RBC lifespan, blood panel data, demographic factors, and uridine diphosphate glucuronosyltransferase (UGT1A1) mutation load were calculated by Spearman analysis. Susceptibility factors for mild hemolysis were analyzed by multivariate regression analysis. RESULTS: The mean RBC lifespan of the GS subjects was significantly shorter than the normal reference value (95.4 ±â€Š28.9 days vs 126 days; t = -7.504, P < .01), with 30.0% below the lower limit of the normal reference range (75 days). The RBC lifespan of the participant with CN2 was 82 days. Serum TB correlated positively with UGT1A1 mutation load (γ = 0.281, P = .048), hemoglobin (γ = .359, P = .010) and hematocrit (γ = 0.365, P = .010), but negatively with RBC lifespan (γ = -0.336, P = .017). No significant susceptibility factors for mild hemolysis were found. CONCLUSIONS: The results indicate that mild hemolysis indeed, exists in a portion of patients with GS and might serve as an important contributor to unconjugated hyperbilirubinemia in addition to UGT1A1 polymorphism. Further studies on the mechanism and the potential risks in various medical treatments might be wanted.

Optimizing the performance of magnet-controlled capsule endoscopy based on radiological and gastroscopic modeling.

Routine use of magnet-controlled capsule endoscopy of the stomach has been limited by the inadequate views of specific stomach regions. In the present study, radiology and upper gastrointestinal endoscopy (UGIE) were used to determine optimal subject body positioning and suitable external control magnet placement for capsule endoscopy. Healthy adult volunteers were subjected to upper gastrointestinal X-ray radiography (n=5), spiral computed tomography with volume reconstruction (n=4) or UGIE (n=1). Stomach fundus-to-body (FB) and body-to-antrum (BA) angles were compared when subjects were supine, prone, lying on their left side and on their right side, and when they were standing upright. Vertical distances from the surface of the body to the distal points of the fundus and antrum were also compared in this range of subject positions. Obtuse angles were considered the most beneficial for capsule movement and short vertical distances were considered desirable for optimizing magnetic force. The FB angle was sharply acute in the supine position, relatively open where subjects were on their side, and almost 180° in the standing position. The BA angle was obtuse in the standing position but acute in all other positions. With the subject in any position, the left lower lateral chest had the shortest distance to the fundus, while the ventral wall was closest to the antrum. The present modeling analysis indicates that standing is superior to all decubitus positions for magnetic-capsule endoscopy, including the commonly used supine position. Both the abdominal anterior wall and left lateral lower chest appeared to be advantageous locations for external control magnet placement.

Human gastric magnet-controlled capsule endoscopy conducted in a standing position: the phase 1 study.

BACKGROUND: Current magnet-controlled capsule endoscopy (MCE) for the stomach is not yet satisfactory with respect to navigation control, especially in the gastric fundus and cardia. A newly developed MCE system conducted in a standing rather than supine position may improve capsule maneuverability within the stomach. The aim of this phase 1 study was to assess the feasibility and safety of this system for examining the human stomach in healthy volunteers. METHODS: A cohort of 31 healthy volunteers were enrolled. Each swallowed a capsule after drinking water and gas producing agents intended to produce distention. Under the newly developed standing MCE system, subjects were examined endoscopically while standing with external guide magnets placed on the abdominal wall and left lower chest. Safety, gastric preparation, maneuverability, visualization of anatomical landmarks and the gastric mucosa, and examination time were the primary parameters assessed. The gastric preparation and examination procedures were well accepted by the subjects and there were no adverse events. RESULTS: Gastric examination took 27.8 ± 8.3 min (12-45 min). Gastric cleanliness was good in 24 participants (77.4%) and moderate in 7 participants (22.6%). Gastric distention was good in all of 31 participants (100%). Capsule maneuverability was also graded as good in all 31 subjects (100%), and manipulation in the fundus and cardia regions was as easy as that in the antrum and body. Visualization of the gastric cardia, fundus, body, angulus, antrum and pylorus was assessed subjectively as complete in all 31 subjects (100%). Visualization of the gastric mucosa was also good (> 75%) in all 31 subjects (100%). In areas where the mucosa could not be visualized, the low visibility was due to opaque fluid or foam. Polyps and erosive lesions were found in 25 subjects. CONCLUSION: MCE of the stomach conducted in a standing position is feasible and safe with satisfactory maneuverability.

Red Blood Cell Lifespan Shortening in Patients with Early-Stage Chronic Kidney Disease.

BACKGROUND: Although reduced red blood cell (RBC) lifespan has been reported to be a contributory factor to anemia in patients with end-stage chronic kidney disease (CKD), there are limited data regarding RBC lifespan in early-stage CKD. Serum erythropoietin (EPO) is considered a primary causative factor of renal anemia. The aims of this study were to compare the RBC lifespan, serum EPO levels, and other renal anemia indicators across CKD-stage groups of patients and to analyze the impacts of etiological factors on renal anemia. METHODS: A cohort of 74 non-smoking patients with CKD were enrolled, including 15 in stage 1, 18 in stage 2, 15 in stage 3, 15 in stage 4, and 11 in stage 5. RBC lifespan was determined by CO breath tests. Potential correlations of hemoglobin (Hb) concentration with RBC lifespan, reticulocyte count (Ret), and levels of EPO, ferritin, folic acid, and vitamin B12 were analyzed. RESULTS: CKD progression was associated with decreases in (Hb) and RBC lifespan. RBC lifespan durations in CKD stages 1-5 were 122 ± 50, 112 ± 26, 90 ± 32, 88 ± 28, and 60 ± 24 days, respectively. RBC lifespan means for the stage 3, 4 and 5 groups were significantly shorter than those for the stage 1 and 2 groups. Serum EPO did not differ significantly between the CKD stage groups. (Hb) correlated directly with RBC lifespan (r = 0.372, p = 0.002) and Ret (r = 0.308, p = 0.011), but did not correlate with serum EPO, ferritin, folic acid, or vitamin B12 levels. CONCLUSIONS: Reduced RBC lifespan in early-stage CKD, demonstrated in this study, suggests that increased RBC destruction may play a more important etiological role in renal anemia than other indicators in patients with CKD.

Effect of Hemodialysis on the Red Blood Cell Life Span in Patients with End-Stage Kidney Disease.

The aim of this study was to use a CO breath test to investigate hemodialysis effects on red blood cell lifespan in patients with chronic kidney disease. A cohort of 17 non-smoking men with end-stage kidney disease undergoing hemodialysis via a polysulfone dialysis membrane (as opposed to a traditional cellulose acetate membrane) were subjected to a repeated Levitt's CO breath test to compare red blood cell lifespan before vs. after dialysis. None of the patients showed significant fluctuations in endogenous CO concentration during the dialysis procedure. The mean red blood cell lifespan was 66.0 ± 31.0 days before dialysis and 72.0 ± 26.0 days after dialysis, with no significant difference between the assessment time points (P > 0.05). In conclusion, dialysis using a polysulfone membrane did not appear to disrupt red blood cells or reduce their lifespan in patients with end-stage kidney disease.

Human erythrocyte lifespan measured by Levitt's CO breath test with newly developed automatic instrument.

Existing standard techniques for erythrocyte (RBC) lifespan measurement, such as quantitation of labeling with isotopes or biotin, are cumbersome and time-consuming. Given that endogenous CO originates mainly from degraded RBCs, a team lead by Levitt developed a CO breath test to enable more efficient RBC lifespan estimation. The purpose of this study was to evaluate the reliability of Levitt's CO breath test method with our newly developed automatic instrument. RBC lifespan measurements conducted by Levitt's CO breath test method were conducted in 109 healthy subjects and 91 patients with chronic hemolytic anemia. In healthy subjects, the RBC lifespan was 126 ± 26 days, similar to values obtained with classical standard labeling methods. RBC lifespan did not differ significantly between males and females or between juveniles and adults, and did not correlate with age. To our knowledge, this datum represents an RBC lifespan average for the largest sample to date. In subjects with hemolytic anemia, RBC lifespan was 29 ± 14 days, which is significantly shorter than that of the healthy subjects (p = 0.001). Using 75 days as a cut-off, diagnostic accuracy for hemolytic anemia in the present study sample was 100%. In conclusion, the present results indicate that Levitt's CO breath test is an ideal method for human RBC lifespan measurement, and the newly developed automatic instrument is reliable and convenient for clinical practice.

Gastric Residual Volume after Split-Dose Bowel Preparation versus Conventional Single-Dose Regimen before Anesthetic Colonoscopy.

The aim of this study was to compare gastric residual volume (GRV) in patients given a split-dose versus a conventional single-dose of polyethylene glycol (PEG) preparation before undergoing anesthetic colonoscopy. Methods. In a prospective observational study, we assessed GRV in outpatients undergoing same-day anesthetic gastroscopy and colonoscopy between October 8 and December 30 of 2016. Outpatients were assigned to the split-dose (1 L PEG in the prior evening and 1 L PEG 2-4 h before endoscopy) or single-dose (ingestion of 2 L PEG ≥ 6 h before endoscopy) regimen randomly. Bowel cleansing quality was assessed with the Boston Bowel Preparation Scale (BBPS). Results. The median GRV in the split-dose group (17 ml, with a range of 0-50 ml; N = 65) was significantly lower than that in the single-dose group (22 ml, with a range of 0-62 ml; N = 64; p = 0.005), with a better bowel cleansing quality (BBPS score 8.05 ± 0.82 versus 7.64 ± 1.21; p = 0.028). GRV was not associated with diabetes or the use of medications. Conclusions. GRV after a split-dose preparation and fasting for 2-4 hours is not larger than that after a conventional single-dose preparation and fasting for 6-8 hours. The data indicates that the split-dose bowel preparation might not increase the risk of aspiration.

Rapid CO breath test screening of drugs for protective effects on ribavirin-induced hemolysis in a rabbit model: a pilot study.

Hemolytic anemia is a major side effect of ribavirin antiviral treatment for chronic hepatitis C. Ribavirin dose reduction may compromise the antiviral response and erythropoietin can take several weeks to alleviate anemia. The purpose of the present study was to screen potentially protective drugs against ribavirin-induced hemolytic anemia in a rabbit model, using our modified CO breath test for measuring erythrocyte (RBC) lifespan, the gold standard diagnostic index of hemolysis. Fifteen rabbits were divided randomly into five groups (N = 3/group): one vehicle control group, one ribavirin (only)-treated (RBV) group, and three groups initially treated with ribavirin only, followed by a combination of ribavirin with prednisone (RBV + Pred), polyene phosphatidyl choline (RBV + PPC), or reduced glutathione (RBV + GSH). RBC lifespan was calculated from accumulated CO measured in a closed rebreath apparatus, blood volume measured by the Evan's blue dye (EBD) dilution test, and hemoglobin concentration data. The RBC lifespan was normal in the vehicle control group (44-60 d), but reduced significantly in all of the ribavirin-treated groups before the addition of screened drugs (17-35 d). RBC lifespan rebounded significantly with the addition of glutathione, but not with the addition of prednisone or polyene phosphatidyl choline. A similar overall drug effect pattern was seen in the hemoglobin concentration and reticulocyte count data. In conclusion, the results of this pilot study indicate that reduced glutathione can attenuate ribavirin-induced hemolytic anemia, and that the RBC lifespan measured with our modified rapid CO breath test is feasible and reliable for use in animal studies.

A Modified Carbon Monoxide Breath Test for Measuring Erythrocyte Lifespan in Small Animals.

This study was to develop a CO breath test for RBC lifespan estimation of small animals. The ribavirin induced hemolysis rabbit models were placed individually in a closed rebreath cage and air samples were collected for measurement of CO concentration. RBC lifespan was calculated from accumulated CO, blood volume, and hemoglobin concentration data. RBC lifespan was determined in the same animals with the standard biotin-labeling method. RBC lifespan data obtained by the CO breath test method for control (CON, 49.0 ± 5.9 d) rabbits, rabbits given 10 mg/kg·d(-1) of ribavirin (RIB10, 31.0 ± 4.0 d), and rabbits given 20 mg/kg·d(-1) of ribavirin (RIB20, 25.0 ± 2.9 d) were statistically similar (all p > 0.05) to and linearly correlated (r = 0.96, p < 0.01) with the RBC lifespan data obtained for the same rabbits by the standard biotin-labeling method (CON, 51.0 ± 2.7 d; RIB10, 33.0 ± 1.3 d; and RIB20, 27.0 ± 0.8 d). The CO breath test method takes less than 3 h to complete, whereas the standard method requires at least several weeks. In conclusion, the CO breath test method provides a simple and rapid means of estimating RBC lifespan and is feasible for use with small animal models.

Pilot validation of the Boston Bowel Preparation Scale in China.

BACKGROUND AND AIM: The Boston Bowel Preparation Scale (BBPS) is a novel bowel cleanliness rating scale that has undergone validation at Boston University Medical Center, Boston, MA, USA. Thus far, there is no standard recognized bowel preparation scale in China. The aim of the present study was to analyze the reliability and validity of the BBPS for the assessment of bowel preparation quality (BPQ) in China. METHODS: A group of 49 participants from several hospitals in Guangdong province viewed a video demonstration of BBPS provided by Boston Medical Center and participated in a continuing education seminar. Inter-observer reliability was assessed for three testing colonoscopies in the video. Three months later, 13 of the participants repeated the test, and intra-observer reliability was assessed. The BBPS was then applied prospectively in 1012 screening colonoscopies and BBPS scores were compared with polyp-detection rate. Intraclass correlation coefficients (ICC) and weighted Kappa values assessed inter- and intra-rater reliability, respectively. The association of BBPS scores with polyp-detection rates was calculated by χ(2) tests. RESULTS: The inter-observer ICC of BBPS scores was 0.987 (95% CI, 0.949-1.0). The weighted Kappa for BBPS scores was 0.671 (95%CI, 0.507-0.841). For 1012 screening colonoscopies, the mean BBPS score was 6.9 ± 1.8. BBPS scores ≥ 5 were associated with a higher polyp-detection rate (35%) than scores < 5 (18%) (P < 0.05). CONCLUSION: The BBPS is a valid and reliable measure of BPQ, and this validity and reliability was maintained for Chinese physicians taught via video.

A Retrospective Analysis to Validate the Alarm Signs Used in the CEDAP-Plus Study.

Background and Study Aim. This study aimed to validate the alarm signs used in the 2007 German CEDAP-Plus study for indicating capsule endoscopy in patients who have idiopathic chronic abdominal pain. Patients and Methods. We retrospectively reviewed the cases of all patients who underwent capsule endoscopy at our institution between August 2007 and August 2009 for chronic hitherto undiagnosed abdominal pain, despite previous investigations. The demographic data, indications, findings, and diagnoses were recorded, as were the alarm signs (i.e., 10% loss of weight within 3 months, suspected small intestinal bleed or chronic anemia, and laboratory indications of inflammation). Results. Alarm signs were found in only 4 of the 62 included patients. Capsule endoscopy revealed findings that led to diagnoses of Crohn's disease (n = 4), tuberculosis (n = 1), gastrointestinal stromal tumors (n = 3), and hookworm (n = 1); these diagnoses included 100% (4/4) of the patients with alarm signs, but only 8.6% (5/58) of patients without them. However, 55.6% (5/9) of patients with clinically capsule endoscopy findings reported no alarm signs. Conclusions. Although selecting patients based on the alarm signs may increase the yield of capsule endoscopy, the alarm sign criteria appear to have low sensitivity.

[The imprinting status of genetic imprinted gene PEG10 in human hepatocellular carcinomas].

OBJECTIVE: To study the imprinting status of genetic imprinted gene PEG10 (perternally expressed gene 10) in human hepatocellular carcinoma (HCC) tissues and liver cancer cell lines. METHODS: Genomic DNA was extracted from 20 HCC tissues and its adjacent tissues, 15 normal liver tissues, 5 liver cancer cell lines (PLC/PRF/5, smmc-7721, HepG2, Hep3B, SK-HEP-1) and 2 normal human liver cell lines (changliver, HL7702). The DNA fragments containing single nucleotide polymorphism (SNP) site of PEG10 were amplified by polymerase chain reaction (PCR) and the genotype of samples was detected by DNA sequencing. Total RNA was extracted from heterozygous samples, the imprinting status and expression level of PEG10 were evaluated by quantitative real time reverse transcription-polymerase chain reaction (qRT-PCR) and DNA sequencing. RESULTS: It was found that 16/40 of HCC and its adjacent tissues were heterozygous, 3/15 of normal liver tissue were heterozygous. A site of heterozygous mutation was found in HepG2 cells by DNA sequencing. The other liver tissues and cell lines were all homozygous. PEG10 was biallelically expressed and showed loss of imprinting (LOI) in 82.4% (14/17) liver cancer samples (16 HCC tissues and HepG2), however it was a monoallelic expression and showed genomic imprinting in17.6% (3/17) liver cancer samples. In HCC, the expression levels of PEG10 were increased apparently, but it was negative expressed in cancer adjacent tissues and normal liver tissues. Expression levels of PEG10 were not significantly different between imprinted HCC tissues and HCC tissues with LOI (t = 1.311, P value is more than 0.05). CONCLUSION: PEG10 imprinting is lost in a majority of HCC and no correlation exists between the imprinting status of PEG10 and its expressions in HCC tissues.

Artesunate induces oncosis-like cell death in vitro and has antitumor activity against pancreatic cancer xenografts in vivo.

Pancreatic cancer is highly resistant to the currently available chemotherapeutic agents. Less than 5% of patients diagnosed with this disease could survive beyond 5 years. Thus, there is an urgent need for the development of novel, efficacious drugs that can treat pancreatic cancer. Herein we report the identification of artesunate (ART), a derivative of artemisinin, as a potent and selective antitumor agent against human pancreatic cancer cells in vitro and in vivo. ART exhibits selective cytotoxic activity against Panc-1, BxPC-3 and CFPAC-1 pancreatic cancer cells with IC(50) values that are 2.3- to 24-fold less than that of the normal human hepatic cells (HL-7702). The pan caspase inhibitor zVAD-fmk did not inhibit the cytotoxic activity of ART. Electron microscopy of ART-treated cells revealed severe cytoplasmic swelling and vacuolization, swollen and internally disorganized mitochondria, dilation (but not fragmentation) of the nuclei without chromatin condensation, and cell lysis, yielding a morphotype that is typical of oncosis. The ART-treated cells exhibited a loss of mitochondrial membrane potential (DeltaPsim) and ART-induced cell death was inhibited in the presence of the reactive oxygen species (ROS) scavenger N-acetyl-cysteine (NAC). Importantly, ART produced a dose-dependent tumor regression in an in vivo pancreatic cancer xenografts model. The in vivo antitumor activity of ART was similar to that of gemcitabine. Taken together, our study suggests that ART exhibits antitumor activity against human pancreatic cancer via a novel form of oncosis-like cell death, and that ART should be considered a potential therapeutic candidate for treating pancreatic cancer.