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Background:. Colorectal cancer (CRC) is the fourth cause of cancer death in China. We aimed to provide national and subnational estimates and changes of CRC premature mortality burden during 2005–2020. Methods:. Data from multi-source on the basis of the national surveillance mortality system were used to estimate mortality and years of life lost (YLL) of CRC in the Chinese population during 2005–2020. Estimates were generated and compared for 31 provincial-level administrative divisions in China. Results:. Estimated CRC deaths increased from 111.41 thousand in 2005 to 178.02 thousand in 2020; age-standardized mortality rate decreased from 10.01 per 100,000 in 2005 to 9.68 per 100,000 in 2020. Substantial reduction in CRC premature mortality burden, as measured by age-standardized YLL rate, was observed with a reduction of 10.20% nationwide. Marked differences were observed in the geographical patterns of provincial units, and they appeared to be obvious in areas with higher economic development. Population aging was the dominant driver which contributed to the increase in CRC deaths, followed by population growth and age-specific mortality change. Conclusions:. Substantial discrepancies were observed in the premature mortality burden of CRC across China. Targeted considerations were needed to promote a healthy lifestyle, expand cost-effective CRC early screening and diagnosis, and improve medical treatment to reduce CRC mortality among high-risk populations and regions with inadequate healthcare resources.

Urinary tract infections (UTIs) are bacterial infections that 1) commonly affect females, 2) can pose high risks to impair kidney function, 3) are often treated with broad-spectrum antibiotics, and 4) are associated with high recurrence rates due to the evolution of drug-resistant strains. To choose the appropriate antibiotic, the minimum inhibitory concentration (MIC) among a panel of antibiotics should be determined before administration to avoid inadequate dosing or use of wrong antibiotics. To meet with the unmet needs, we developed a bead-based method for bacterial preconcentration with capture rates ranging from 20-50% and then automatically performed on-chip AST on an automated device which was composed of a pneumatic control module, a temperature control module and a chip image processing module. The developed portable system was capable of automatically conducting AST and MIC measurements using urine samples (via image analysis) in only 4.5-9 h and tested on four common UTIs bacterial strains. This compact system may therefore be promising for point-of-care personalized medicine in the near future.

Aims: Catheter ablation of paroxysmal atrial fibrillation (PAF) has been shown to be effective and safe. However, recurrence of PAF varies between 10 and 30% for radiofrequency ablation. There have been no reports comparing long-term recurrence rates following radiofrequency ablation, cryoablation, and three-dimensional guided cryoablation plus radiofrequency ablation. The aim of this study was to observe the long-term recurrence rate of PAF when treated by these three catheter ablation methods, and to explore clinical factors that can potentially predict PAF recurrence following catheter ablation.Methods: There were 238 patients involved in this study, including 106 radiofrequency (RF) ablation cases (RF group), 66 cryoablation cases (Freeze group), and 66 cases treated by three-dimensional guided cryoablation combined with radiofrequency ablation (Freeze-plus-RF group). All patients underwent standardized follow-up. The recurrence rate of atrial fibrillation (AF) in the three groups was calculated. Predictive factors for the recurrence of AF were also investigated.Results: At 48 months (the median follow-up period), the sinus rhythm maintenance rate was 77.4% in the RF group, 72.7% in the Freeze group, and 81.8% in the Freeze-plus-RF group. The maintenance rate of sinus rhythm was highest in the Freeze-plus-RF group, but differences among the three groups were not statistically significant. Further analysis found that the preoperative left atrial appendage emptying velocity (LAAEV) (recurrence vs. no recurrence, 56.58 ± 18.37 vs. 65.59 ± 18.83, respectively, p = 0.003), left atrial (LA) anteroposterior dimension (recurrence vs. no recurrence, 36.56 ± 4.65 vs. 35.00 ± 4.37, respectively; p = 0.028), and LA vertical dimension (recurrence vs. no recurrence, 56.31 ± 6.96 vs. 53.72 ± 6.52, respectively; p = 0.035) were related to postoperative recurrence. Multiple Cox regression analysis showed that only LAAEV was predictive of postoperative recurrence of PAF (hazard ratio: 0.979; 95% confidence interval: 0.961–0.997).Conclusion: Our study found that there was no statistically significant difference in long-term recurrence rates among the RF, Freeze, and Freeze-plus-RF groups. Preoperative LAAEV is an independent predictor of postoperative recurrence of PAF.

Purpose Adult-onset immunodeficiency with anti-interferon (IFN)-γ autoantibodies (anti-IFN-γ Abs) is an immunodeficiency syndrome. Immune dysfunction in this distinct disorder remains to be clarified. Methods We prospectively collected blood samples of 20 patients with anti-IFN-γ Abs and 65 healthy normal subjects. Percentages of lymphocyte subpopulations, most relevant to T-, B-, and NK-cells, and percentages of stimulated lymphocytes with cytokine production were assessed using eight-color flow cytometry. The results were adjusted to age and absolute lymphocyte counts.Results Most (85%) patients presented non-tuberculous mycobacterial infection. Skin lesions were predominantly manifested by neutrophilic dermatoses. Involved lymph nodes had granulomatous inflammation, except 22.2% showing atypical lymphoid hyperplasia without granuloma formation. In the multiple linear regression model, CD4+ T cells and non-activated subpopulations (recent thymic emigrants and naïve subtypes) were significantly decreased with increased expression of activation markers and polarization to Th1, Th17, and Treg. The percentage of NK cells was increased, but two major NK subpopulations, CD56bright and CD56+CD16+ subsets were decreased. Furthermore, NK cells diminished expression of NKp30 and NKp46 with increased CD57 expression. The cytokine production was significantly lower, namely TNF-α in CD4+ T cells (P = 0.009), CD8+ T cells (P < 0.001), and NK cells (P = 0.002); IFN-γ in CD8+ T cells (P = 0.002) and NK cells (P = 0.001); and IL-2 in CD4+ (P < 0.001), and CD8+ (P = 0.005) T cells. Conclusion We conclude that the immune system in patients with anti–IFN-γ Abs could be exhausted which may contribute to the distinct clinicopathologic features.

Background Catheter ablation has emerged as a major strategy for paroxysmal atrial fibrillation (PAF). Atrial electrical remodeling (AER) plays a critical role in the recurrence of PAF after ablation. Hypothesis To characterize the immediate trends of AER during ablations in patients with PAF, and assess the relationship between immediate trends and recurrence. Methods We performed this prospective observational study of 135 patients to investigate AER following three ablation modes: radiofrequency ablation (RFA), cryoablation (CA) and 3D mapping‐guided cryoablation (3D‐CA). The atrial effective refractory period (AERP) and atrial conduction time (ACT) were measured via electrophysiology before and immediately after ablation, and P‐wave indices were measured via electrocardiography before and within 24 h after ablation. Follow‐up visits were conducted for at least 1 year or until relapse. Results Different approaches of ablation caused a fairly significant increase in the shortest P‐wave duration and AERP in both the proximal coronary sinus (PCS) and distal coronary sinus (DCS) but caused a shortened P‐wave dispersion. No different effect was found at the AERP among the three modes. Compared to patients who received CA, among patients who received RFA, a significant reduction in total ACT and right ACT was seen. Statistically, there was a weakly positive association between changes in total ACT and early recurrence. Conclusions Injury during ablation for PAF was associated with an increase in the AERP but not in the ACT. Total ACT and right ACT were shorter after RFA than after CA. The increase in total ACT were slightly predictive of early recurrence.

Background: A single procedure combining left atrial appendage closure (LAAC) plus catheter ablation (CA) has been proven to be safe and feasible for treating atrial fibrillation (AF). However, the influence of treatment modality on left atrial (LA) function is not thoroughly explained.Objective: We aimed to investigate the changes of LA function in persistent AF patients undergoing concomitant LAAC and CA.Methods: The study population comprised 65 patients who underwent combined AF ablation and Watchman LAAC (combined therapy group) in our center, and 65 participants of the AF simple ablation group who were matched based on sex, age, CHA2DS2-VASc score and HAS-BLED score using propensity score matching. During the 1-year follow-up period, two-dimensional echocardiography and speckle tracking echocardiography were performed to assess LA reservoir, conduit, and contractile function.Results: The combined therapy was associated with a significant improvement in the LA reservoir function with increased expansion index and strain indices, including strain and strain rate (SR) during ventricular systole. Conduit function with SR during early ventricular diastole was also improved, as was contractile function with active atrial emptying fraction and SR during atrial systole. Similarly, LA reservoir and contractile function indices all improved continuously during follow-up after catheter ablation alone. At 3 months follow-up LA reservoir and conduit function with strain indices had a tendency to improve only in the simple procedure group. At 1-year follow-up there was no significant difference in either LA volumes or strain indices between the two groups.Conclusion: Both the combined therapy group and the simple ablation group demonstrated significant improvement in LA function. Based upon the fact that LA function was improved in both groups it might be concluded that most of the effects appeared to result from ablation, not LAAC; furthermore the additional LAAC procedure did not affect the improvement of LA function after CA.

CUL4A regulate the termination of autophagy in a physical process. However, the relationship between CUL4A and autophagy in cancer is unclear. We retrospectively investigated 99 intrahepatic cholangiocarcinoma (iCCA) cases. Whole sections were used for immunohistochemical analysis for p62, and LC3B expression. Q-score was defined as the sum of the labeling intensity and proportion. The cut-off point for immunoreactivity was set. CUL4A was overexpressed in cell lines and autophagy reflux was compared after manipulation. The iCCA cases with CUL4A overexpression had significantly higher prevalence of intact activated autophagy (42.4 vs. 15.2%; p = 0.003), which was significantly associated with advance tumor stage (34.1% vs. 15.4%; p = 0.032), less extensive necrosis (8.3 vs. 49.3%; p < 0.001), and shortened disease-free survival (mean, 19.6 vs. 65.5 months, p = 0.015). In vitro, iCCA cells with CUL4A overexpression significantly increased LC3II level as compared to the cells under basal condition. Although both cell types showed intact autophagy with increased LC3II expression after bafilomycin A1 treatment, the accumulation of LC3II was higher in CUL4A-overexpressing cells. CUL4A overexpression increased the proliferation of cells as compared with control cells. After treatment with bafilomycin A1, proliferation was inhibited in both cell types, but the effects were more prominent in the cells overexpressing CUL4A. CUL4A promotes autophagy, and exhibits significantly higher autophagic flux which affects the proliferation of iCCA cells; these effects correlated with advance tumor stage and poor prognosis. Thus, targeting autophagy may be potentially therapeutic in iCCA.

Primary brainstem haemorrhage (PBH) is characterized by acute onset, rapid deterioration, many complications, and poor prognosis. Its treatment has been controversial. This study aimed to explore the clinical risk factors of postoperative survival and neurological function recovery of stereotactic aspiration in the treatment of PBH. The clinical data of 65 patients with severe brainstem haemorrhage from February 2019 to February 2020 in the First Hospital of Hebei Medical University were reviewed. All patients were treated with stereotactic haematoma aspiration. We determined the survival status of patients at 30 days after the operation and the recovery of neurological function at 90 days. The modified Rankin Scale score (mRS) was used to assess the survival status. The 30-day mortality rate was 23.1% (15 patients). The proportion of patients with good neurological recovery at 90 days after the operation was 32.3% (21 patients). According to the multivariate logistic regression analysis, the haematoma classification was an independent risk factor for postoperative survival (OR = 0.197, 95% CI: 0.016–0.385, p = 0.046) and recovery of neurological function 90 days after surgery (OR = 0.019, 95% CI: 0.001–0.267, p = 0.003). The haematoma classification is an independent risk factor for 30-day mortality and recovery of neurological function 90 days after surgery. Massive and basal-tegmental haematomas were associated with higher mortality. The prognosis of patients with unilateral and bilateral tegmental haematoma was better than that of patients with other haematoma types.

Since early diagnosis of sepsis may assist clinicians in initiating timely, effective, and prognosis-improving antibiotic therapy, we developed an integrated microfluidic chip (IMC) for rapid isolation of both Gram-positive and Gram-negative bacteria from blood. The device comprised a membrane-based filtration module (90 min operating time), a bacteria-capturing module using a micro-mixer containing magnetic beads coated with "flexible neck" regions of mannose-binding lectin proteins for bacteria capture (20 min), and a miniature polymerase chain reaction (PCR) module for bacteria identification (90 min via TaqMan® probe technology). The filter separated all white blood cells and 99.5% of red blood cells from bacteria, which were captured at rates approaching 85%. The PCR assay's limit of detection was 5 colony-forming units (CFU) per reaction, and the entire process was completed in only 4 h. Since this is far less than that for culture-based approaches, this IMC may serve as a promising device for detection of sepsis.

This study reports an integrated microfluidic device that was capable of executing rapid antimicrobial susceptibility tests with one, two, or even three antibiotics against two clinically isolated multi-drug-resistant bacteria strains (including carbapenem-resistant Escherichia coli and methicillin-resistant Staphylococcus aureus). Bacteria were automatically mixed for 10 min with serially diluted antibiotics with a novel, membrane-type micromixer consisting of two circular micropumps, and the minimum inhibitory concentrations (MIC) were then determined via simple colorimetric reactions in only 4.5–6 h using only 3 μL of bacteria sample of each reaction (as opposed to 24 h and 50 μL, respectively, with the conventional broth micro-dilution method). In addition to determining MICs of antibiotics (ceftazidime, gentamicin, meropenem, vancomycin and linezolid), interaction effects across antibiotics combinations (gentamicin/meropenem or ceftazidime/gentamicin/meropenem) at different dosages were explored. The efficacy of polypharmacy showed additivity when gentamicin or ceftazidime/gentamicin were combined with meropenem to treat carbapenem-resistant Escherichia coli. This represents the first time that the perplexing clinical decision to choose multiple antibiotics for combination therapy against drug resistant bacteria can be realized on an integrated microfluidic device within 6 h.

Objectives Severe hyperkalemia can cause life-threatening arrhythmia, cardiac arrest, or death. This study aimed to investigate the incidence and the associated factors relevant to critical hyperkalemia (≥6 mmol/L) among inpatients, outpatients, and emergency department. Their clinical outcomes were also analyzed. Methods All patients whose high serum potassium values had been reported as critical laboratory values in 2016 were enrolled. Their demographic data, comorbidities, clinical symptoms, biochemical data, and outcomes were reviewed and collected. The Charlson comorbidity score (CCS) and glomerular filtration rate (GFR) were computed to assess the comorbidity burden and renal function. Patients were divided into groups according to different settings, potassium and GFR levels, and their survival. Results Of the 293,830 total serum potassium tests, 1,382 (0.47%) reports were listed as critical laboratory values. The average reply time was 6.3 min. Their mean age was 67.2 years, while the average GFR was 12.2 mL/min/1.73 m2. The overall mortality rate was 34%. Patients in the emergency department had the highest incidence (0.92%), while inpatients had the worst outcome (51% mortality). The leading cause of mortality was septic shock. The fatal group had higher rates of clinical symptoms, higher potassium values, CCS, and eGFR (all p Conclusions Most of the responses for the reports were obtained within a short period of time. Patients with reported high critical serum potassium values were characterized by high rates of comorbidity, reduced eGFR, and mortality. The incidence, clinical manifestations, and outcomes varied in the different clinical settings.

Background: Catheter ablation for atrial fibrillation (AF) has been gaining popularity; however, the trend of inflammatory response markers in patients treated with different catheter ablation strategies over time and their predictability of AF recurrence remain unknown. Methods: A total of 210 patients with AF were enrolled and grouped according to surgical mode as follows: freeze group, RF group, and freeze3D group. The subjects were tested for related indexes before and after surgery. To determine AF recurrence during follow up, 24-h ambulatory electrocardiography was performed at two, three, six, and 12 months after surgery. Results: The inflammation indexes of the three groups peaked between one and three days after surgery but fell at different time points (P < .05). The recurrence rate of paroxysmal atrial fibrillation (PAF) was positively correlated with the increase in the percentage of white blood cells and neutrophils after surgery (P < .05). Conclusions: The postoperative inflammation indices peaked and fell at different time points after different catheter ablation methods. In addition, the recurrence rate of AF in patients treated with freeze3D is lower.

The current coronavirus disease 2019 (COVID-19) pandemic has caused significant challenges throughout the world and a rapid, reliable diagnostic test is in high demand. Real-time reverse transcription polymerase chain reaction (RT-PCR) was one of the most quickly established methods of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection and is considered to be the gold standard. In this report, we share our experience of using two different testing platforms: the cobas 6800 SARS-CoV-2 test, an automated system that was recently granted Emergency Use Authorization by the FDA, and a laboratory-developed test based on the protocol from the Taiwan Centers for Disease Control (CDC). There was an overall 96.2% agreement between the two platforms. However, the positive agreement between the two platforms was only 80.0%. We found 3 instances of discordance between the two systems and this emphasized the need for timely diagnosis with a reliable testing platform.

Background The influenza virus is a highly infectious disease, with a notably rapid transmission rate. Autophagy is triggered by viral infection and is a survival mechanism exerted to maintain cellular homeostasis. Catechin is a representative phenolic acid which exerts anti-inflammatory responses against influenza A virus infection. The aim of this study is to explore the anti-H1N1 influenza virus effects by catechin associated with the restoration of autophagy. Methods XTT assay was used to detect cellular viability. The inhibitory effects on the H1N1 influenza virus were assessed by hemagglutination assay, neuraminidase activity, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The protein levels of H1N1 influenza virulence and autophagic markers were detected by Western blot. Results We herein demonstrated that catechin had no cytotoxic effect on both infected and noninfected A549 cells and exerted protective effects on infected A549 cells. The results of the hemagglutination assay, neuraminidase activity, and qRT-PCR to examine viral load demonstrated that catechin effectively inhibited the replication of the H1N1 influenza virus. The virulent M2 protein and viral nucleoprotein were also inhibited after treatment with catechin. As for the autophagic markers, the LC3B protein was notably decreased by catechin in a dose-dependent manner, while the amount of autophagic vacuoles in H1N1 influenza virus-infected cells also decreased after catechin treatment in a dose-dependent manner. Conclusion Collectively, the autophagy activated by the H1N1 influenza virus could be reversed after catechin treatment. This study indicates that catechin effectively inhibits H1N1 viral proliferation and thus may be applied as an adjuvant in future clinical application.

Objective: The present study was to compare the incidence of septal defect (SD) in patients with atrial fibrillation (AF) who received radiofrequency ablation or cryoablation. Methods: A total of 293 AF patients were performed with radiofrequency ablation and cryoablation. Cardiac ultrasonography was performed to calculate left atrial diameter (LAD), left atrial ejection fraction (LAEF%), strain rate (SR), left ventricular systolic (SRs), left ventricular diastolic (SRe), and left atrial systole (SRa) before surgery, 3 months and 1 year after surgery. The patients were followed up to observe statin and angiotensin-converting enzyme inhibitor (ACEI)/angiotensin receptor blocker (ARB) medication, AF recurrence, 6-min walk test, stroke, any symptoms caused by arrhythmia, and re-hospitalization. Results: The levels of LAD and SD were higher, while SRe and SRa were lower in the cryoablation group in the comparison with the radiofrequency ablation group after surgery (P

Rapid diagnosis of sepsis assists clinicians to initiate timely antibiotic therapy and therefore reduce mortality of bacterial infections. In this work, we presented an integrated microfluidic system for fast isolation of bacteria for both gram-positive and gram-negative bacteria directly from human whole blood, which will be crucial in the detection of sepsis. First, a deterministic lateral displacement (DLD) device was used for sample pretreatment of whole blood, which performed continuous separation of leukocytes, erythrocytes and bacteria. With this approach, most of the blood cells were removed while bacteria and platelets were collected. Then the isolated bacteria were captured with magnetic beads surface-coated with flexible neck regions of mannose-binding lectin (FcMBL) using a new 3D micromixer. The entire process including sample pretreatment and bacteria isolation could be completed on an integrated microfluidic system within 2 hours, which is significantly less than the one for the traditional method (i.e. 3 days). This is the first time that an integrated microfluidic system capable of detecting bacteria by using FcMBL was reported, which may provide a promising way to diagnose sepsis.

Ferritin is a ubiquitous and conserved iron storage protein that plays a significant role in host detoxification, iron storage, and immune response. Although ferritin has been studied in many species, little is known about its role in the Eri-silkworm (Samia cynthia ricini). In this study, the ferritin light-chain subunit gene, named ScFerLCH, was identified from S. c. ricini. The full-length gene, ScFerLCH, was 1,155 bp and encoded a protein consisting of 231 amino acids with a deduced molecular weight of 26.38 kDa. Higher ScFerLCH expression levels were found in the midgut, silk gland, and fat body by quantitative reverse-transcription polymerase chain reaction and western blot analysis. Injection of Staphylococcus aureus and Pseudomonas aeruginosa could induce upregulation of ScFerLCH in the hemolymph, fat body, and midgut, indicating that ScFerLCH may contribute to the host defense against invading pathogens. In addition, the native ferritin protein was isolated from S. c. ricini by native polyacrylamide gel electrophoresis and its two subunits, ferritin heavy-chain subunit (ScFerHCH) and ferritin light-chain subunit (ScFerLCH), were identified by mass spectrometry. Specifically, we found that recombinant ferritin subunits could self-assemble into a protein complex in vitro; moreover, both recombinant subunits and the protein complex were found to bind different bacteria, including Escherichia coli, P. aeruginosa, S. aureus, and Bacillus subtilis. However, bactericidal tests showed that the protein complex could not inhibit the growth of bacteria directly. Taken together, our results suggest that ScFerritin might play an important role in mediating molecular interaction with pathogens.

Low-level cadmium exposure has adverse effects on chronic kidney disease (CKD); however, the risk factors for elevated blood cadmium levels (BCLs) have not been studied in CKD. We conducted a cross-sectional investigation in 200 CKD patients and stratified them by the tertiles of BCL to compare their demographic, environmental, and biochemical data. The factors associated with BCL were identified, and their effects were examined in subgroups. In the analyses, female sex, smoking, and CKD stage 5D were associated with high BCL, and statin was inversely correlated with BCL (odds ratio [95% confidence interval, CI], 6.858 [2.381–19.746], p < 0.001, 11.719 [2.843–48.296], p = 0.001, 30.333 [2.252–408.520], p = 0.010, and 0.326 [0.122–0.873], p = 0.026; deviations of BCL [nmol/L, 95% CI], 2.66 [1.33–4.00], p < 0.001, 3.68 [1.81–5.56], p < 0.001, 3.38 [0.95–5.82], p = 0.007, and −2.07 [−3.35–−0.78], p = 0.002). These factors were also independently correlated with BCL in subgroups, including non-dialysis CKD, hypertensive patients, non-smokers, and male patients. In conclusion, female sex, smoking, and CKD stage 5D were the major risk factors for elevated BCL; additionally, statins were negatively associated with BCL in CKD.

Bombyx mori nucleopolyhedrovirus (BmNPV) is a serious pathogenic microorganism that causes tremendous loss to sericulture. Previous studies have found that some proteins of serine protease family in the digestive juice of B. mori larvae have anti-BmNPV activity. In our previous publication about proteome analysis of the digestive juice of B. mori larvae, the digestive enzyme trypsin, alkaline A (BmTA) was filtered as a differentially expressed protein possibly involved in BmNPV resistance. Here, the biological characteristics and anti-BmNPV functions of BmTA were comprehensively analysed. The cDNA sequence of BmTA had an ORF of 768 nucleotides encoding 255 amino acid residues. Domain architecture analysis showed that BmTA contained a signal peptide and a typical Tryp_SPc domain. Quantitative real-time PCR analysis showed that BmTA was highly expressed in the larval stages and specifically expressed in the midgut of B. mori larvae. The expression level of BmTA in BmNPV resistant strain A35 was higher than that in susceptible strain P50. After BmNPV infection, the expression of BmTA increased in both strains from 24 to 72 h. Virus amplification analysis showed that the relative levels of VP39 in B. mori larvae and BmN cells infected with the appropriate concentration of recombinant-BmTA-treated BmNPV were significantly lower than in the control groups. Moreover, overexpression of BmTA in BmN cells significantly inhibited the amplification of BmNPV. Taken together, the results of this study indicated that BmTA possessed anti-BmNPV activity in B. mori, which broadens the horizon for virus-resistant breeding of silkworms.

Background CUL4A has been known for its oncogenic properties in various human cancers. However, its role in intrahepatic cholangiocarcinoma (iCCA) has not been explored. Methods We retrospectively investigated 105 iCCA cases from a single medical institution. Tissue microarrays were used for immunohistochemical analysis of CUL4A expression. CUL4A expression vectors were introduced in cell lines. Cell migration and invasion assays were used to compare the mobility potential of iCCA cells under basal conditions and after manipulation. Then we evaluated the effects of CUL4A on the cell growth by proliferation assay, and further checked the susceptibility to cisplatin in iCCA cells with or without CUL4A overexpression. Results CUL4A overexpression was detected in 34 cases (32.4%). Patients with CUL4A-overexpressing tumors exhibited shortened disease-free survival (mean, 27.7 versus 90.4 months; P = 0.011). In the multivariate analysis model, CUL4A overexpression was shown to be an independent unfavorable predictor for disease-free survival (P = 0.045). Moreover, stably transfected CUL4A-overexpressing iCCA cell lines displayed an increased mobility potential and enhanced cell growth without impact on susceptibility to cisplatin. Conclusions Our data demonstrate that overexpression of CUL4A plays an oncogenic role in iCCA and adversely affects disease-free survival. Thus, it may prove to be a powerful prognostic factor and a potential therapeutic target.

Exposure to phthalates has been demonstrated to cause reproductive toxicity in animals, but evidence of the association between phthalates and markers of male reproductive function have been inconsistent in human studies. Here we examined whether environmental exposure to phthalates contributes to altered reproductive hormone levels, sperm DNA damage and spermatozoa apoptosis in a Chinese population. From March to June 2013, repeated urine samples collected from male partners of couples attending an infertility clinic in Wuhan, China were analyzed for 8 phthalate metabolites. Associations of the urinary phthalate metabolites with serum hormone levels (n=483), sperm DNA damage parameters (n=509) and spermatozoa apoptosis measures (n=467) were assessed using multivariable linear regression models. After adjusting for potential confounders, mono-(2-ethylhexyl) phthalate (MEHP), a metabolite of di-(2-ethylhexyl)-phthalate (DEHP), was inversely associated with serum levels of estradiol, total testosterone (T) and free T (all P for trend

Atrial fibrillation (AF) is a commonly encountered heart arrhythmia and a risk factor for cardiovascular system. The purpose of the present study was to explore the role of long non-coding RNA myocardial infarction-associated transcript (MIAT) in AF and AF-induced myocardial fibrosis and the possible mechanisms involved in this process. We successfully induced an AF rat model. Expression of MIAT presented a dramatic increase, while microRNA (miR)-133a-3p presented a dramatic decrease in atrium tissues of rats with AF induction. In addition, we also found that MIAT was highly expressed and miR-133a-3p was significantly reduced in peripheral blood leukocyte of AF patients. For biological function exploration of MIAT/miR-133a-3p axis, MIAT was knocked down using small hairpin RNA (shRNA) lentivirus injection and the rescue experiments were performed simultaneously by inhibiting miR-133a-3p using anti-miR-133a-3p lentivirus injection in rats with AF. MIAT downregulation significantly alleviated AF, increased atrial effective refractory period (AERP), and reduced the duration of AF as well as cardiomyocytes apoptosis. Whereas these effects of MIAT downregulation on AF were reversed by anti-miR-133a-3p administration. Luciferase reporter revealed that miR-133a-3p was directly regulated by MIAT. Moreover, MIAT knockdown effectively reduced AF-induced atrial fibrosis by detecting reduced collagen in the right atria and inhibited expression of fibrosis-related gene expression of collagen I, collagen III, connective tissue growth factor (CTGF) and transforming growth factor-β1 (TGF-β1) in rats with AF, these findings were in contrast with the findings for rats with inhibition of miR-133a-3p. In conclusion, our study demonstrated the role of MIAT downregulation in alleviating AF and AF-induced myocardial fibrosis, and the functional regulatory pathway of MIAT targeting miR-133a-3p.

Polypharmacy is routinely administered to fight severe infections, though it has led to rampant multi-drug resistance in many bacterial strains. Preferably, antimicrobial susceptibility testing (AST) would be carried out prior to antibiotic prescription, though it is generally thought to be too complex and labor-intensive. In order to assist clinicians with better antibiotic administration for the effective treatment of bacterial infections, an integrated microfluidic system (IMS) capable of automating AST for 1-2 antibiotics against clinical bacterial pathogens was developed herein. Accurate determination of the minimum and fractional inhibitory concentrations of vancomycin, gentamicin, and linezolid were determined by assaying growth of two clinical methicillin-resistant Staphylococcus aureus isolates via a colorimetric assay on-chip. By applying various antibiotic combinations against a single pathogen in multiple chambers, the IMS could identify the optimal drug combination and the minimum effective dosage by evaluating the fractional inhibitory concentration index. This IMS possessed several advantages over conventional methods, including (1) a 50% reduction in bacterial sample and reagent volume (

Fungal keratitis (FK) is one of the most severe corneal infectious diseases. FK often leads to poor visual prognosis and thus requires accurate diagnosis. Conventional approaches, including clinical diagnoses, smears, and cultures, often fail to provide reliable diagnostic value. Omics approaches, such as those using genomic, metagenomic, and tear proteomic data sources, provide promising features for improving the diagnosis and monitoring the progression of FK. Genomic approaches are based mainly on detecting amplicons of ribosomal RNA genes, and internal transcribed spacers are gradually gaining popularity in clinical practices. A metagenomic approach based on 16S rRNA genes may help monitor the dynamic change of conjunctival microbiota associated with an FK event, whereas that based on shot-gun and 18S rRNA target enrichment sequencing could have the potential to diagnose FK using clinical samples. A tear proteomic approach may provide comprehensive information about ocular surface defense and injury during FK. Representative up- and down-regulated proteins during FK could also be used as biomarkers to determine the clinical course and develop a treatment strategy in different stages of FK. Consequently, a personalized tear proteomic approach will soon play a key role in FK management.

This study reported a portable microfluidic system which can be used to automatically perform antibiotic screening with single antibiotic, double antibiotic combinations and even triple antibiotic combinations on clinically isolated bacteria strains. With the incorporation of the integrated pneumatically-driven microfluidic chip, a pneumatic control module, a temperature control module, and an optical detection module, the developed system was capable of achieving significant improvement of operation flexibility as well as cost reduction for determination of antimicrobial susceptibility test and the efficacy of drug combinations via colorimetric results within 4 hours. Personalized antibiotic screening may be realized by using the developed microfluidic system for rapid clinical decision-making.

Restoration of sinus rhythm in atrial fibrillation (AF) by radiofrequency catheter ablation (RFCA) is associated with a transient stunning of left atrial (LA) function. However, the long-term effects of different ablation strategies on LA function remain undetermined. We performed randomized controlled trial to evaluate the effects of RFCA, cryoablation, and 3D mapping-guided cryoablation on LA function of proximal AF patients within 1 year. The 3D mapping-guided cryoablation was defined as a maximum of two cryoablation procedures for each pulmonary vein accompanied by RFCA for additional points until complete pulmonary vein isolation was achieved. Conventional and speckle tracking echocardiographic analyses were performed to evaluate LA function. Among the 210 patients (70 in each group) included, a trend of decreasing LA systolic and diastolic function was observed in all groups, as evidenced by decreases in peak A-wave velocity, the global LA peak systolic strain, the peak strain rate, the peak early diastolic strain rate, and the peak late diastolic strain rate within 7 days to 3 months after ablation followed by gradual recovery thereafter. However, the temporal changes in the above four strain parameters among the three groups did not differ significantly within 1 year after ablation (all p > 0.05). Parameters of the LA emptying fraction and LA dimensions were not significantly affected. These results suggested that stunning of LA function occurred within 7 days to 3 months after ablation, and different strategies of AF ablation did not differentially affect the temporal changes in LA function up to 1 year after ablation.

Background/objectives Docosahexaenoic acid (DHA), an n-3 long chain polyunsaturated fatty acid (LCPUFA), is acquired by dietary intake or the in vivo conversion of α-linolenic acid. Many enzymes participating in LCPUFA synthesis are regulated by peroxisome proliferator-activated receptor alpha (PPARα). Therefore, it was hypothesized that the tissue accretion of endogenously synthesized DHA could be modified by PPARα. Materials/methods The tissue DHA concentrations and mRNA levels of genes participating in DHA biosynthesis were compared among PPARα homozygous (KO), heterozygous (HZ), and wild type (WT) mice (Exp I), and between WT mice treated with clofibrate (PPARα agonist) or those not treated (Exp II). In ExpII, the expression levels of the proteins associated with DHA function in the brain cortex and retina were also measured. An n3-PUFA depleted/replenished regimen was applied to mitigate the confounding effects of maternal DHA. Results PPARα ablation reduced the hepatic Acox, Fads1, and Fads2 mRNA levels, as well as the DHA concentration in the liver, but not in the brain cortex. In contrast, PPARα activation increased hepatic Acox, Fads1, Fads2 and Elovl5 mRNA levels, but reduced the DHA concentrations in the liver, retina, and phospholipid of brain cortex, and decreased mRNA and protein levels of the brain-derived neurotrophic factor in brain cortex. Conclusions LCPUFA enzyme expression was altered by PPARα. Either PPARα deficiency or activation-decreased tissue DHA concentration is a stimulus for further studies to determine the functional significance.

Campylobacter spp. may cause fever, vomiting, and diarrhea in humans. Antibiotic treatment is suggested for patients with severe campylobacteriosis. However, the interpretative criteria for antibiotic susceptibility are inconsistent between Clinical and Laboratory Standards Institute and European Committee on Antimicrobial Susceptibility Testing guidelines. The aim of the study is to investigate the antibiotic susceptibility and prevalence of cytolethal distending toxin genes and to evaluate antibiotic susceptibility testing methods in the clinical laboratories of two tertiary medical centers in Taiwan. In total, 236 bacterial isolates were collected between 2001 and 2014. The disk diffusion and E-test methods were used to evaluate the antibiotic susceptibility, and broth dilution results were used as a reference. The virulence genes cdtA, cdtB, cdtC, and ceuE were detected through polymerase chain reaction. The antimicrobial sensitivity rates for erythromycin, ciprofloxacin, and tetracycline using the broth dilution assay were 80.4, 5.4, and 3.4%, respectively. No significant differences were observed in the antibiotic susceptibility of the isolates obtained from southern and northern Taiwan. However, some differences were observed between species. The susceptibility test for erythromycin (disk diffusion) showed that the isolates with small inhibition zone diameters were all resistant, and five isolates (4.0%) with large IZDs were non-sensitive. The error rate of the disk diffusion method according to the CLSI M45-A3 guideline was 5.4% (8/148). The incompatibility rates between the E-test and broth dilution methods for erythromycin, ciprofloxacin, and tetracycline were 8.0, 5.3, and 1.3%, respectively. The positive rates of the genes cdtA and cdtC were considerably higher in Campylobacter jejuni than in C. coli. Erythromycin is recommended as the first choice of treatment for campylobacteriosis. The disk diffusion method is suitable for prescreening Campylobacter susceptibility by using the CLSI M45-A2 and EUCAST criteria (low error rate of 3.4%). If antibiotic treatment fails or IZDs are between 6 and 20 mm, minimum inhibitory concentration testing by using the E-test method is highly recommended because the results of the E-test and broth dilution methods exhibit high agreement. The error rate of disk diffusion method using CLSI M45-A3 criteria is only slightly higher than B, which is also a suitable criteria.

Periprosthetic joint infections (PJIs) are among the most severe complications emerging from prosthetic joint replacement surgeries. In order to possess a rapid means of diagnosing PJIs, an integrated microfluidic system was developed herein for detecting and identifying bacteria in human synovial fluid (HSF). The entire molecular diagnostic process, including (1) sample treatment, (2) bacterial isolation, (3) bacterial lysis, (4) nucleic acid amplification (via polymerase chain reaction (PCR)), and (5) optical detection, could be automated on a single chip. First, N-acetyl-L-cysteine was used to decrease the viscosity of HSF samples and consequently enhance bacterial isolation with vancomycin-coated nano-magnetic beads. Then, a universal 16S ribosomal ribonucleic acid PCR primer set and four species-specific primer sets were used for PCR-based detection and identification of four common bacteria previously associated with PJIs, including Staphylococcus aureus, methicillin-resistant S. aureus, Escherichia coli, and Acinetobacter baumannii. With this approach, the limit of detection was as low as 100 colony forming units (CFUs) per milliliter (or 20 CFUs per reaction), which is suitable for clinical diagnostics and for making informed decisions regarding post-operative antibiotic administration. More importantly, bacterial detection and identification data could be acquired within 90 minutes, representing a significant improvement over traditional culture-based methods (3–7 days). The developed microfluidic system may therefore serve as a promising tool for rapid diagnosis of PJIs.

Background Neurofibromatosis type 1 (NF1) is one of the most common hereditary neurocutaneous disorders. The malignant peripheral nerve sheath tumor (MPNST), transformed from NF1 related plexiform neurofibroma, is a rapidly growing and highly invasive tumor. No effective chemotherapeutic agent is currently available. Calebin-A is a derivative from turmeric Curcuma longa. Given the anti-inflammatory and anticancer potentials of curcumin, whether Calebin-A also had the tumoricidal effect upon MPNST cells is still elusive. Purpose To determine whether Calebin-A has the potential for anti-MPNST effect. Methods The MTT and FACS analysis of normal Schwann (HSC) and MPNST cells have been employed to determine the tumoricidal effect of Calebin-A. The expression of the signal pathway molecules was assessed by Western blotting. The CHIP with quantitative PCR assay was performed to quantify the promoter DNA binding to acetylated histone 3 (acetyl H3). The enzyme activities of histone acetyltransferase (HAT) and deacetylase (HDAC) have been evaluated by commercial kits. The measurements of tumor size of the xenograft mouse model were also performed. Results Calebin-A inhibited the proliferation of MPNST and primary neurofibroma cells in a dose-dependent manner. The flow cytometry analysis of the MPNST cells after treatment of 25 μm of Calebin-A demonstrated an increase of population in the G0/G1 phase but decrease in G2/M phase. Before treatment, the expression of Axl, Tyro3, and acetyl H3 was significantly higher in MPNST cells when compared to HSC. The expression of phosphorylated-AKT, -ERK1/2, survivin, hTERT, and acetyl H3 proteins were reduced after treatment. The CHIP assay shows the promoter DNA copies of survivin (BRIC5) and hTERT genes are significantly reduced post-treatment. The enzyme activity of HAT was significantly reduced, but not that of HDAC. Two HAT inhibitors, epigallocatechin-3-gallate (EGCG) and anacardic acid (AA) have also demonstrated a significant inhibitory effect on MPNST cells. Finally, the measurements of tumor size showed a significant reduction of the xenograft tumors after treatment of Calebin-A. Conclusion Both in vitro and in vivo studies showed Calebin-A could inhibit the proliferation of MPNST with suppression of survivin and hTERT. The reduced expression of these two factors might be through the epigenetic histone modification resulting from the decreased activity of HAT.

Aims The aim of this study was to compare the results of 16S/28S rRNA sequencing with the erythrocyte sedimentation rate (ESR), C-reactive protein (CRP) level, and synovial fluid analysis in the diagnosis of prosthetic joint infection (PJI). Patients and Methods Between September 2015 and August 2016, 214 consecutive patients were enrolled. In the study population, there were 25 patients with a PJI and 189 controls. Of the PJI patients, 14 (56%) were women, and the mean age at the time of diagnosis was 65 years (38 to 83). The ESR and CRP levels were measured, and synovial fluid specimens were collected prospectively. Synovial fluid was subjected to reverse transcription polymerase chain reaction (RT-PCR)/sequence analysis targeting the 16S/28S rRNA, and to conventional culture. Laboratory personnel who were blind to the clinical information performed all tests. The diagnosis of PJI was based on the criteria of the Musculoskeletal Infection Society. Results A total of 25 patients had a confirmed PJI. In 20 cases of monomicrobial PJI, the PCR products could be perfectly matched with the 16S/28S rRNA genes specific for different species of bacteria provided by sequence analysis. Of the five polymicrobial cases of PJI, 16S/28S rRNA PCR sequence analysis failed to identify the concordant bacteria species. In the 189 control patients, there was one false-positive RT-PCR result. The sensitivity and specificity of the molecular diagnosis method were 100% (95% confidence interval (CI) 85.7 to 100) and 99.5% (95% CI 97.1 to 99.9), respectively, whereas the positive and negative predictive values of PCR were 96.1% (95% CI 79.6 to 99.9) and 100% (95% CI 98.1 to 100), respectively. The PCR results were significantly better than serological diagnostic methods (p = 0.004 and p = 0.010 for ESR and CRP, respectively), the synovial fluid white blood cell (WBC) count (p = 0.036), and percentage of polymorphonuclear cells (PMN%) (p = 0.014). Conclusion Stepwise RT-PCR and sequence analysis of the 16S/28S rRNA carried out under stringent laboratory conditions achieved highly sensitive and specific results for the differentiation between aseptic and septic joints undergoing arthroplasty. Sequence analysis successfully identified bacterial strains in monomicrobial infections but failed to identify molecular targets in polymicrobial infections. Further refinement of the protocols to identify the bacteria in polymicrobial infections is needed. Cite this article: Bone Joint J 2018;100-B:1345–51.

Extranodal nasal-type natural killer (NK)/T-cell lymphoma (NKTCL) is an aggressive lymphoma that is prevalent among East Asian and South American populations. Although Epstein-Barr virus (EBV) is commonly detected in NKTCL, there are limited studies that have analyzed the EBV genomic variations in NKTCL. In this study, 8 EBV latent genes were analyzed using targeted gene sequencing in 23 formalin-fixed paraffin-embedded tissues derived from 18 patients with NKTCL. Five cases with paired samples were comparatively analyzed. The consistency of EBV sequencing data between tissue samples was high (96.3%–98.7%), whereas that of variant calling among the tissue samples and plasma samples (74.3%–79.2%) was low. The highest densities of non-synonymous variants were detected in the EBNA3B gene. Among the 74 known T-cell epitopes, 363 non-synonymous variants were identified in 32 (43.2%) epitopes. Additionally, the AVFDRKSDAK (A1S/P and V2F/M/L) and YHLIVDTDSL (I4L and L10R/V/G/H) epitopes were associated with 5 patterns of amino acid changes in EBNA3B and EBNA-2, respectively. The frequency of variation in the human leukocyte antigen (HLA)-restricted epitopes with corresponding HLA types common among Taiwanese population was significantly low (P = 0.011), whereas that in anchor residues was significantly high (P = 0.012). In conclusion, this study demonstrated the genomic diversity of EBV in NKTCL and its correlation with the HLA-restricted epitope variations in Taiwanese population. The findings of this study provide useful insights for the development of novel therapeutic strategies for NKTCL.

Genome-wide association studies (GWAS) have identified Aldehyde dehydrogenase 2 (ALDH2) as a susceptibility locus for coronary artery disease (CAD) previously. However, the impacts of common variants in this gene on CAD and its outcomes have not been extensively studied. This study explored the association between the Tagging SNPs in ALDH2 and CAD as well as its main outcomes. Six common variants in ALDH2 were selected as tagging SNPs and two cohorts containing 7296 individuals were genotyped to investigate the impacts of ALDH2 on CAD and its main outcomes. The results show that the variant rs671 in ALDH2 is associated with an increased risk of CAD in southern Chinese (OR=1.26, 95%CI: 1.07-1.48, p=0.004), while not in northern Chinese (OR=1.00, 95%CI: 0.86-1.50, p=0.94). Meanwhile, we find that rs671 genotypes may not influence the outcomes of CAD (HR=1.11, 95%CI: 0.892-1.38, p=0.346). Additionally, we also tested the effect of rs671 genotype on CAD severity, while no significant association was found between them. In the subgroup analysis, the results revealed that rs671 were significantly associated with CAD (OR=1.24, 95%CI: 1.11-1.38, p

Exposure to phthalates has been found to have adverse effects on male reproductive function in animals. However, the findings from human studies are inconsistent. Here we examined the associations of phthalate exposure with semen quality and reproductive hormones in a Chinese population using phthalate metabolite concentrations measured in semen as biomarkers. Semen (n = 687) and blood samples (n = 342) were collected from the male partners of sub-fertile couples who presented to the Reproductive Center of Tongji Hospital in Wuhan, China. Semen quality parameters and serum reproductive hormone levels were determined. Semen concentrations of 8 phthalate metabolites were assessed using high-performance liquid chromatography and tandem mass spectrometry. Associations of the semen phthalate metabolites with semen quality parameters and serum reproductive hormones were assessed using confounder-adjusted linear and logistic regression models. Semen phthalate metabolites were significantly associated with decreases in semen volume [mono-n-butyl phthalate (MBP), mono-(2-ethylhexyl) phthalate (MEHP), mono(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono(2-ethyl-5-oxohexyl) phthalate (MEOHP)], sperm curvilinear velocity [monobenzyl phthalate (MBzP), MEHP, the percentage of di-(2-ethylhexyl)-phthalate metabolites excreted as MEHP (%MEHP)], and straight-line velocity (MBzP, MEHP, %MEHP), and also associated with an increased percentage of abnormal heads and tails (MBzP) (all p for trend

Exposure to phthalates has been demonstrated to have adverse effects on male reproduction in animal studies, but findings in human studies have been inconsistent. We recruited 1040 men from the Reproductive Center of Tongji Hospital in Wuhan, China from March to June 2013. Each man provided one semen sample and two urine samples. Semen quality parameters and the urinary concentrations of eight phthalate metabolites were determined. After multivariable adjustments, the urinary concentrations of monobutyl phthalate (MBP) were found to be positively associated with the below-reference sperm concentration and total sperm count, and the odds ratios (ORs) comparing extreme MBP quartiles were 2.01 (95% CI: 1.07, 3.79; p for trend=0.06) and 1.80 (95% CI: 1.05, 3.08; p for trend=0.02), respectively. The associations were confirmed by multivariable linear regression analysis, which showed that the MBP concentration was significantly associated with decreasing trends in the sperm concentration and total sperm count (both p for trend

Objective To investigate myocardial apoptosis and the expression of apoptosis-related proteins [B-cell lymphoma-2 (Bcl-2) and Bax], cardiac hypertrophy, and function in spontaneously hypertensive rats (SHR) during aging. Methods Male SHR (n = 36) and Wistar-Kyoto rats (WKY, n = 30) were monitored for their cardiac function from 14 to 102 weeks of age using echocardiography. Myocardial apoptosis in the subendocardium and subepicardium was analyzed using TUNEL staining. The apoptosis-related proteins (Bcl-2 and Bax) were assessed by Western blot analysis. Results Left ventricular mass index (LVMI) in SHR increased progressively with age. Left ventricular ejection fraction (LVEF) showed no significant difference from 14 to 66 weeks of age, but decreased significantly in SHR from 84 to 102 weeks of age. The apoptotic index (APOI) of myocardial cells in SHR increased with age, in which subendocardial APOI (APOI-endo) increased at 66 weeks and was significantly higher than the age-matched WKY (9.83 ± 3.97 vs. 4.03 ± 2.06, P < 0.05). Additionally, the subepicardial APOI (APOI-epi) increased significantly at 84 weeks of age (P < 0.05). Bax/Bcl-2 ratio was higher in SHR than WKY at 66 weeks (3.98 ± 0.80 vs. 0.29 ± 0.06, P < 0.05), and enhanced further at 102 weeks of age (10.54 ± 1.60 vs. 0.70 ± 0.13, P < 0.05). Myocardial APOI was positively correlated with LVMI (r = 0.83, P < 0.01), negatively with LVEF in 66–102 weeks (r = −0.81, P < 0.01), and positively with Bax/Bcl-2 ratio (r = 0.79, P < 0.01). Conclusion Myocardial apoptosis increases with aging and is associated with progressive left ventricular remodeling and cardiac dysfunction in SHR.

Bombyx mori nucleopolyhedrovirus (BmNPV) is a major pathogen causing severe economic loss. Previous studies have revealed that some proteins in silkworm digestive juice show antiviral activity. In this study, antiviral activity examination of different resistant strains showed that the digestive juice of the resistant strain (A35) had higher inhibition to virus than the susceptible strain (P50). Subsequently, the label-free quantitative proteomics was used to study the midgut digestive juice response to BmNPV infection in P50 and A35 strains. A total of 98 proteins were identified, of which 80 were differentially expressed proteins (DEPs) with 54 enzymes and 26 nonenzymatic proteins by comparing the proteomes of infected and non-infected P50 and A35 silkworms. These DEPs are mainly involved in metabolism, proteolysis, neuroactive ligand receptor interaction, starch and sucrose metabolism and glutathione metabolism. After removing the genetic background and individual immune stress response proteins, 9 DEPs were identified potentially involved in resistance to BmNPV. Further studies showed that a serine protease, an alkaline phosphatase and serine protease inhibitor 2 isoform X1 were differentially expressed in A35 compared to P50 or post BmNPV infection. Taken together, these results provide insights into the potential mechanisms for silkworm digestive juice to provide resistance to BmNPV infection. Signifcance: Bombyx mori nucleopolyhedrovirus (BmNPV) is highly pathogenic, which has a great impact on the sericulture. BmNPV entered the midgut lumen and exposed to digestive juices after oral infection. Previous studies have revealed that some proteins in silkworm digestive juice show antiviral activity, however, current information on the digestive juice proteome of high resistant silkworm strain after BmNPV challenge compared to susceptible strain is incomprehensive. Here, we combined label-free quantification method, bioinformatics, RT-qPCR and western blot analysis and found that BmNPV infection causes some protein changes in the silkworm midgut digestive juice. The DEPs were identified in the digestive juices of different resistant strains following BmNPV infection, and screened out some proteins potentially related to resistance to BmNPV. Three important differentially expression proteins were validated by independent approaches. These findings uncover the potential role of silkworm digestive juice in providing resistance to BmNPV and supplemented the profile of the proteome of the digestive juices in B. mori., Graphical abstract Unlabelled Image, Highlights • The digestive juice antiviral activity of resistant strain was higher than susceptible strain. • Silkworm digestive juice was analyzed by label-free quantitative proteomics. • Eighty DEPs were identified in different resistant strains following BmNPV infection. • Some DEPs were validated by RT-qPCR and western blot. • These results provide insights into the potential mechanisms for B. mori resistant to BmNPV infection.

Background and Purpose: Drynaria fortunei J. Sm (D. fortunei), known as Gu-Sui-Bu, is used in traditional Chinese medicine to treat common injuries, including bone fractures and bruising. The specific functional mechanisms of the angiogenic and endothelial cell migration properties of D. fortunei are currently unclear. Thus, the purpose of this study is to validate the potential angiogenic and cellular migration properties and related mechanisms by D. fortunei both in vivo and in vitro. Experimental Approach: The present study investigates, both in vivo and in vitro, the wound healing effects of D. fortunei as associated with angiogenesis, specifically by the modulation of matrix metalloproteinases (MMPs) and upregulation of vascular endothelial growth factor (VEGF) ligand/receptors. In order to determine the potential angiogenic effects of D. fortunei, in vivo neovascularization of chick chorioallantoic membranes (CAMs) assay, and directed in vivo angiogenesis assay (DIVVA) were performed, while in vitro scratch wound healing, migration, and matrix-induced tube formation assays were performed by using human umbilical vascular endothelial cells (HUVECs). Furthermore, we used qPCR to analyze the gene expressions and Western blot to observe protein expressions of MMP-2, MMP-14, TIMP-2, RECK, and VEGF/VEGFRs. Results: This study identified five major compounds from the water extract of D. fortunei: protocatechuic acid, caffeic acid 4-O-β-D-glucopyranoside, 5,7-dihydroxychromone-7-O-rutinoside, neoeriocitrin, and naringin. D. fortunei was confirmed to activate in vivo angiogenesis by CAM and DIVVA assays. D. fortunei further exhibited in vitro angiogenic effects associated with cell migration, as demonstrated by the tube formation assay, transwell migration assay, and scratch wound healing assay. The extracellular MMP-2 activity was found to be dose-dependently augmented both in vitro and in vivo by D. fortunei. The mRNA and protein expressions of MMP-2, and MMP-14 were increased; while the tissue inhibitor metalloproteinase-2 (TIMP-2), and reversion-inducing cysteine-rich protein with kazal motifs (RECK) were both decreased. Furthermore, D. fortunei activated the gene and protein expressions of VEGF-A, -B, and VEGFR-2, -3. Conclusion: D. fortunei increased MMP-2 activity, thereby stimulating angiogenesis and cell migration, both in vivo and in vitro, as a result of MMP-2 and TIMP-2 balance modulation and the activation of VEGF/VEGFRs expression.

Carnosic acid (CA), a major polyphenolic diterpene present in Rosmarinus officinalis, has been reported to have multiple functions, including antitumor activity. The MTT assay, BrdU incorporation, wound healing, and colony formation were used to detect melanoma B16F10 cell growth and proliferation. Flow cytometry was used for cell cycle detection. p21 and p27 expression was detected by Western blotting. B16F10 cell xenograft model was established, and treated with CA, carmustine (BCNU), or lomustine (CCNU). The present study found that CA exhibits significant growth inhibition and cell cycle arrest in melanoma B16F10 cells. We also found that CA triggers cell cycle arrest at G0/G1 phase, and enhances p21 expression. Additionally, CA can enhance BCNU- and CCNU-mediated cytotoxicity and cell cycle arrest in B16F10 cells. Finally, we found that CA inhibits tumor growth, and reduces the values of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in vivo. The present study study concluded that CA may be safe and useful as a novel chemotherapeutic agent.

We sought to investigate the association of single nucleotide polymorphisms (SNPs) of the genes involved in βAR signaling with the response of patients to βAR blockers. A total of 2403 hospitalized patients with chronic heart failure (HF) were enrolled in a multicenter observational study as the first cohort and followed up for a mean period of 20 months. Genes for β1AR, β2AR, and the major cardiac G-protein-coupled receptor kinases (GRKs) GRK2 and GRK5 were analyzed to identify SNPs, and patients were stratified according to genotypes. A second independent cohort enrolling 919 patients with chronic HF was applied to validate the observed associations. The signaling properties of the key identified SNPs were assessed in vitro. Our data showed that HF patients harboring the Gly16 allele in the gene for β2AR (ADRB2) had an increased risk of the composite end point relative to patients who were homozygous for Arg16. Notably, these patients showed a beneficial response to βAR-blocker treatment in a G allele-dose-dependent manner, whereas Arg16 homozygotes had no response to βAR-blocker therapy. This Arg16Gly genotype-dependent heterogeneity in clinical outcomes of HF was successfully validated in the second independent population. Besides, the in vitro experiments revealed that G allele carriers were defective in β2AR-coupled inhibitory adenylate cyclase g (Gi) protein signaling.

The World Health Organization, in a 2016 revision, introduced recurrent genetic abnormalities for classifying mature T- and NK-cell neoplasms. However, the role of genetic analyses from lymph node aspiration cytology is still not elucidated. We hypothesize that the use of genetic analyses may increase the accuracy of diagnosis from cytological preparations.Fifty-seven formalin-fixed paraffin-embedded (FFPE) samples were collected for next-generation sequencing (NGS) targeting potential driver mutations including TET2, DMN3TA, IDH2, RHOA, STAT3, and STAT5B. Competitive allele-specific TaqMan polymerase chain reaction (cast-PCR) was performed to validate the mutation status by using FFPE and preoperative fine needle aspiration cytology (FNAC) samples.Among these six candidate genes, only IDH2 and RHOA mutations were significantly more frequent in nodal subtypes, angioimmunoblastic T-cell lymphoma (AITL) and peripheral T-cell lymphoma, not otherwise specified (PTCL, NOS) (P = .002 and0.001, respectively). All genes exhibited different mutation patterns except RHOA with a hotspot mutation involving the Gly17 residue. The RHOA G17V mutation was found in 15 (75%) of 20 AITL and two (22%) of nine PTCL, NOS. Cast-PCR using FFPE samples showed 100% concordance with NGS. Among 12 lymph node aspirates, the preliminary diagnoses were suspicious for lymphoma (3, 25%), atypical lymphoid cells (3, 25%) and benign/negative (6, 50%). Cast-PCR detected the RHOA G17V mutation in six (75%) of eight RHOA-mutated aspirates and revealed negative results in all (100%) of four wild-type aspirates, with an 83.3% (10/12) concordance comparing to FFPE samples.The RHOA G17V mutation serves as a useful biomarker for cytological assessment in AITL. The use of cast-PCR is valuable in the diagnosis of malignant lymphomas from cytological preparations, and thus avoiding the potential risks of invasive procedures.

Limited data are available about the role of common variants at the aldehyde dehydrogenase 2 gene (ALDH2) on the clinical outcome in Chinese patients with coronary heart disease (CHD) undergoing percutaneous coronary intervention (PCI). In the present study, a total of 1089 patients were consecutively enrolled from January 2012 and July 2013. Six common variants at ALDH2 gene, including rs2339840, rs4648328, rs4767939, rs11066028, rs16941669, and rs671, were selected to test the associations of those polymorphisms with the cardiovascular outcome in patients with CHD after PCI. The clinical endpoints included cardiovascular death, nonfatal myocardial infarction, and nonfatal stroke. The composite of clinical endpoints was defined as the primary endpoint, and every endpoint alone was considered as the secondary endpoints. The median follow-up time was 38.27 months. Our results showed that the common variant rs2339840 was independently associated with a lower risk of stroke in patients with CHD after PCI (codominant model, HR = 0.32, 95% CI, 0.11–0.91, P = .074 for heterozygotes; HR = 0.25, 95% CI, 0.06–1.14, P = .033 for homozygotes; dominant model, HR = 0.32, 95% CI, 0.14–0.74, P = .007). However, no significant associations were found between other 5 single nucleotide polymorphisms (SNPs) and the clinical endpoints. For the first time, the common variant rs2339840 was reported to be a protective factor against stroke in CHD patients with PCI.

Systematic examination of the inputs and outputs of the nonlemniscal auditory thalamus will facilitate the functional elucidation of this complex structure in the central auditory system. In mice, comprehensive tracing studies that reveal the long-range connectivity of the nonlemniscal auditory thalamus are lacking. To this end, we used Cre-inducible anterograde and monosynaptic retrograde viruses in Calbindin-2A-dgCre-D and Calretinin-IRES-Cre mice, focusing on the differences across subdivisions of the nonlemniscal auditory thalamus. We found that, 1) the dorsal and medial parts of the auditory thalamus were predominantly connected to sensory processing centers, whereas the posterior intralaminar (PIN) and peripeduncular nucleus (PP) were additionally connected to emotion and motivation modulation centers; 2) ventral auditory cortical areas were the major source of cortical inputs for all subdivisions, and the PIN/PP received more inputs from cortical layer 5 than other subdivisions did; 3) deep layers of the superior colliculus and rostral part of the nonlemniscal inferior colliculus preferentially projected to the PIN/PP; and 4) compared with the dorsal auditory thalamus, the PIN/PP mainly innervated association cortices. In addition, new brain areas connected to the nonlemniscal auditory thalamus, mostly the PIN/PP, were identified. Our results suggested subdivision-specific function of the nonlemniscal auditory thalamus in sound processing.

An integrated microfluidic system was developed to identify bacteria in human joint fluid (HJF) in this study. The entire process including bacteria isolation in HJF, bacteria lysis, polymerase chain reaction (PCR) and optical detection could be automated on a single chip. An universal 16S rRNA primer set and four designed primers to identify Staphylococcus aureus (SA), methicillin-resistance Staphylocuccus aureus (MRSA), Escherichia coli (E. coli), and Acinetobacter baumannii $(\pmb{A}.\pmb{B}.)$ were used for bacteria identification. In addition, N-acetyl-L-cysteine for dissolving sticky mucus of HJF samples and vancomycin-coated magnetic beads were applied for bacteria isolation such that the detection limit could be significantly improved. The entire process could be automated by using a microfluidic system. Experimental results showed that the limit of detection (LOD) could be significantly improved to 100 colony formation units (CFU) per milliliter (or 20 CFU/reaction), which meets the demand on clinical diagnosis. Furthermore, the whole process could be performed within 90 minutes, which is much faster when compared to the traditional method (usually 3–7 days).

Peripheral mature T and NK cell lymphomas consist of a heterogeneous group of neoplasms with cytogenetic and molecular diversities. TP53 mutation is involved in the events of tumorigenesis and present in a variety of cancer subtypes. However, TP53 mutation in peripheral mature T and NK cell lymphomas has not been extensively investigated. In this study, 57 formalin-fixed paraffin-embedded samples were collected for whole-exome sequencing and immunohistochemical study. Ten cases had TP53 mutation variant allele frequency (VAF) of more than 5%, predominantly in intestinal T-cell lymphoma (60%), peripheral T-cell lymphoma, not otherwise specified (22.2%), and extranodal nasal-type NK/T-cell lymphoma (21.7%). Six of 16 (37.5%) cases with 72Pro homozygous genotype showed higher frequency of TP53 mutation VAF > 5% than did those with 72Arg homozygous genotype (P = .044). To a variable extent, overexpression of p53 was observed in 11 of 56 (19.6%) tumors. The percentage of tumor cells with strong p53 staining was positively correlated with TP53 mutation VAF (R2 = 0.95, P < .001). TP53 mutation and p53 expression tended to indicate poor prognosis (P = .054 and P = .009, respectively). In conclusion, we demonstrated TP53 mutation and p53 expression in subtypes of peripheral mature T and NK cell lymphomas and found positive correlation of this immunostaing pattern with the status of TP53 mutation.

Acute leukemia is one of the commonly diagnosed neoplasms and causes human death. However, the treatment for acute leukemia is not yet satisfactory. Studies have shown that mushroom-derived polysaccharides display low toxicity and have been used clinically for cancer therapy. Therefore, we set out to evaluate the anti-cancerous efficacy of a water-soluble polysaccharide extract from Inonotus taiwanensis (WSPIS) on human acute monocytic leukemia THP-1 and U937 cell lines in vitro. Under our experimental conditions, WSPIS elicited dose-dependent growth retardation and induced apoptotic cell death. Further analysis showed that WSPIS-induced apoptosis was associated with a mitochondrial apoptotic pathway, such as the disruption of mitochondrial membrane potential (MMP), followed by the activation of caspase-9, caspase-3, and PARP (poly(ADP-ribose) polymerase) cleavage. However, a broad caspase inhibitor, Z-VAD.fmk, could not prevent WSPIS-induced apoptosis. These data imply that mechanism(s) other than caspase might be involved. Thus, the involvement of endonuclease G (endoG), a mediator arbitrating caspase-independent oligonucleosomal DNA fragmentation, was examined. Western blotting demonstrated that WSPIS could elicit nuclear translocation of endoG. MMP disruption after WSPIS treatment was accompanied by intracellular reactive oxygen species (ROS) generation. However, pretreatment with N-acetyl-l-cysteine (NAC) could not attenuate WSPIS-induced apoptosis. In addition, our data also show that WSPIS could inhibit autophagy. Activation of autophagy by rapamycin decreased WSPIS-induced apoptosis and cell death. Taken together, our findings suggest that cell cycle arrest, endonuclease G-mediated apoptosis, and autophagy inhibition contribute to the anti-cancerous effect of WSPIS on human acute monocytic leukemia cells.

Insulin receptor substrate 2 (IRS2) is a candidate driver oncogene frequently amplified in cancer and is positively associated with IRS2 expression. The overexpression of IRS2 has been suggested to promote tumor metastasis. However, its function in intrahepatic cholangiocarcinoma (iCCA) has not been investigated extensively. The present study examined 86 cases of iCCA to analyze IRS2 expression and its correlation with clinicopathological characteristics using immunohistochemical assays. Three stable cell lines overexpressing IRS2 were established. The mobility potential of cells was compared in the basal condition and following manipulation using cell migration and invasion assays. Epithelial-mesenchymal transition (EMT)-associated proteins were assessed by western blotting. IRS2 was overexpressed in 29 iCCA cases (33.7%) and was significantly more frequent in cases with large tumor size (P=0.033), classified as an advanced stage by the American Joint Committee on Cancer (P=0.046). In comparison with the control cells, the three IRS2-overexpressing iCCA cell lines exhibited a statistically significant increase in mobility potential. Expression analysis of EMT markers demonstrated decreased epithelial marker levels and increased mesenchymal marker levels in IRS2-overexpressing cells compared with their corresponding control cells. The results of the present study indicate that IRS2 overexpression is characterized by a large tumor size and advanced tumor stage in iCCA, and that it may increase tumor mobility potential by regulating EMT pathways. Therefore, it is a valuable predictive indicator of metastasis and may provide a novel direction for targeted therapy in iCCA.