Electrochemical Monitoring of Paclitaxel-Induced ROS Release from Mitochondria inside Single Cells.

Mitochondria are believed to be the major source of intracellular reactive oxygen species (ROS). However, in situ, real-time and quantitative monitoring of ROS release from mitochondria that are present in their cytosolic environment remains a great challenge. In this work, a platinized SiC@C nanowire electrode is placed into a single cell for in situ detection of ROS signals from intracellular mitochondria, and antineoplastic agent (paclitaxel) induced ROS production is successfully recorded. Further investigations indicate that complex IV (cytochrome c oxidase, COX) is the principal site for ROS generation, and significantly more ROS are generated from mitochondria in cancer cells than that from normal cells. This work provides an effective approach to directly monitor intracellular mitochondria by nanowire electrodes, and consequently obtains important physiological evidence on antineoplastic agent-induced ROS generation, which will be of great benefit for better understanding of chemotherapy at subcellular levels.

Sensitive analysis of multiple low-molecular-weight thiols in a single human cervical cancer cell by chemical derivatization-liquid chromatography-mass spectrometry.

Low-molecular-weight (LMW) thiols are important small molecules that regulate or maintain redox homeostasis in physiological and pathological processes. Assessing the concentrations of LMW thiols in biological systems may provide valuable information regarding physiological processes and the early diagnosis of some diseases. Here, we developed a method to simultaneously determine the concentrations of multiple LWM thiols in single cells by chemical derivatization assisted liquid chromatography-mass spectrometry (LC-MS). In this method, we synthesized a pair of stable isotope reagents, N-(acridin-9-yl)-2-bromoacetamide (AYBA) and N-(1,2,3,4-[2H4]-acridin-9-yl)-2-bromoacetamide ([2H4]AYBA). AYBA was used to derivatize LWM thiols in human cervical cancer (HeLa) cells, while [2H4]AYBA was used to derivatize standard LWM thiols to prepare internal standards for the LC-MS method development. The proposed AYBA derivatization greatly enhanced the detection sensitivity of LWM thiols by LC-MS, and thereby achieved the simultaneous detection of multiple LWM thiols by LC-MS in ∼1000 HeLa cells. Finally, the developed method was successfully utilized for the quantitative analysis of multiple LWM thiols in a single HeLa cell and the content changes of LWM thiols in a single HeLa cell before and after oxidative stress treatment. Accordingly, six LMW thiols were detected, including cysteamine, cysteine, glutathione, homocysteine, hydrogen sulfide, and pantetheine.

Validation of the kidney failure risk equation for end-stage kidney disease in Southeast Asia.

BACKGROUND: Patients with chronic kidney disease (CKD) are at high risk of end-stage kidney disease (ESKD). The Kidney Failure Risk Equation (KFRE), which predicts ESKD risk among patients with CKD, has not been validated in primary care clinics in Southeast Asia (SEA). Therefore, we aimed to (1) evaluate the performance of existing KFRE equations, (2) recalibrate KFRE for better predictive precision, and (3) identify optimally feasible KFRE thresholds for nephrologist referral and dialysis planning in SEA. METHODS: All patients with CKD visiting nine primary care clinics from 2010 to 2013 in Singapore were included and applied 4-variable KFRE equations incorporating age, sex, estimated glomerular filtration rate (eGFR), and albumin-to-creatinine ratio (ACR). ESKD onset within two and five years were acquired via linkage to the Singapore Renal Registry. A weighted Brier score (the squared difference between observed vs predicted ESKD risks), bias (the median difference between observed vs predicted ESKD risks) and precision (the interquartile range of the bias) were used to select the best-calibrated KFRE equation. RESULTS: The recalibrated KFRE (named Recalibrated Pooled KFRE SEA) performed better than existing and other recalibrated KFRE equations in terms of having a smaller Brier score (square root: 2.8% vs. 4.0-9.3% at 5 years; 2.0% vs. 6.1-9.1% at 2 years), less bias (2.5% vs. 3.3-5.2% at 5 years; 1.8% vs. 3.2-3.6% at 2 years), and improved precision (0.5% vs. 1.7-5.2% at 5 years; 0.5% vs. 3.8-4.2% at 2 years). Area under ROC curve for the Recalibrated Pooled KFRE SEA equations were 0.94 (95% confidence interval [CI]: 0.93 to 0.95) at 5 years and 0.96 (95% CI: 0.95 to 0.97) at 2 years. The optimally feasible KFRE thresholds were > 10-16% for 5-year nephrologist referral and > 45% for 2-year dialysis planning. Using the Recalibrated Pooled KFRE SEA, an estimated 82 and 89% ESKD events were included among 10% of subjects at highest estimated risk of ESKD at 5-year and 2-year, respectively. CONCLUSIONS: The Recalibrated Pooled KFRE SEA performs better than existing KFREs and warrants implementation in primary care settings in SEA.

Electrochemical Monitoring of ROS/RNS Homeostasis Within Individual Phagolysosomes Inside Single Macrophages.

The existence of a homeostatic mechanism regulating reactive oxygen/nitrogen species (ROS/RNS) amounts inside phagolysosomes has been invoked to account for the efficiency of this process but could not be unambiguously documented. Now, intracellular electrochemical analysis with platinized nanowire electrodes (Pt-NWEs) allowed monitoring ROS/RNS effluxes with sub-millisecond resolution from individual phagolysosomes impacting onto the electrode inserted inside a living macrophage. This shows for the first time that the consumption of ROS/RNS by their oxidation at the nanoelectrode surface stimulates the production of significant ROS/RNS amounts inside phagolysosomes. These results establish the existence of the long-postulated ROS/RNS homeostasis and allows its kinetics and efficiency to be quantified. ROS/RNS concentrations may then be maintained at sufficiently high levels for sustaining proper pathogen digestion rates without endangering the macrophage internal structures.

Consumption of Coffee but Not of Other Caffeine-Containing Beverages Reduces the Risk of End-Stage Renal Disease in the Singapore Chinese Health Study.

Background: Cross-sectional studies suggest that coffee drinking is associated with better renal function. However, to our knowledge, no prospective study has examined its relation with the risk of end-stage renal disease (ESRD). Objective: We examined the relations between coffee, tea, soda, and total caffeine consumption and the risk of ESRD among middle-aged and older Chinese in Singapore. Methods: We used data from the Singapore Chinese Health Study, a prospective cohort of 63,257 men and women aged 45-74 y at recruitment from 1993 to 1998. Baseline information on the consumption of caffeinated coffee and other caffeinated beverages (tea and sodas), habitual diet, medical history, and lifestyle factors was obtained via in-person interviews. The standard serving size of 1 cup was assigned as 237 mL in the questionnaire. Incident ESRD cases were identified via linkage with the nationwide registry. We used multivariable Cox regression models to estimate HRs and 95% CIs of ESRD risk associated with the consumption of caffeinated beverages, with adjustment for potential confounders. Results: After a mean follow-up of 16.8 y, 1143 cohort subjects developed ESRD. Compared with those who drank coffee less than daily, the HR (95% CI) was 0.91 (0.79, 1.05) for those who drank 1 cup of coffee/d and 0.82 (0.71, 0.96) for those who drank ≥2 cups/d (P-trend = 0.012). When stratified by sex, this association was observed in men but not in women. Compared with those who drank less than daily, the HR (95% CI) for drinking ≥2 cups/d was 0.71 (0.57, 0.87) among men and 0.97 (0.78, 1.19) among women (P-interaction = 0.03). Conversely, intakes of tea, soda, or total caffeine were not associated with the risk of ESRD in multivariable models. Conclusion: The consumption of ≥2 cups of coffee/d may reduce the risk of ESRD in the general population, especially among men. This study was registered at http://www.clinicaltrials.gov as NCT03356340.

Red Meat Intake and Risk of ESRD.

Randomized controlled trials suggest that protein restriction may retard the progression of CKD toward ESRD. However, the effects of dietary protein intake level and the food sources of dietary protein on the risk of ESRD in the general population remain unclear. We investigated these effects in the Singapore Chinese Health Study, a prospective population-based cohort that recruited 63,257 Chinese adults aged 45-74 years from 1993 to 1998. We collected habitual diet information via a validated semiquantitative food frequency questionnaire and identified ESRD via record linkage with a nationwide registry. In all, 951 cases of ESRD occurred over a mean follow-up of 15.5 years. Regarding total protein intake, compared with the lowest quartile, the three higher quartiles combined had a hazard ratio for ESRD of 1.24 (95% confidence interval [95% CI], 1.05 to 1.46), but the dose-dependent association across the quartiles was not statistically significant (Ptrend=0.16). Red meat intake strongly associated with ESRD risk in a dose-dependent manner (hazard ratio for highest quartile versus lowest quartile,1.40 [95% CI, 1.15 to 1.71; Ptrend<0.001]). Intake of poultry, fish, eggs, or dairy products did not associate with risk of ESRD. In substitution analysis, replacing one serving of red meat with other food sources of protein associated with a maximum relative risk reduction of 62.4% (95% CI, 33.1 to 78.9; P<0.01). Our study shows that red meat intake may increase the risk of ESRD in the general population and substituting alternative sources of protein may reduce the incidence of ESRD.

Increased body mass index is a risk factor for end-stage renal disease in the Chinese Singapore population.

The relationship between body mass index (BMI) and end-stage renal disease (ESRD) is confounded by co-morbidities associated with both conditions. Furthermore, the association at low range BMI is controversial. We studied this association in the Singapore Chinese Health Study, a population-based prospective cohort that recruited Singaporean Chinese men and women 45-74 years of age from 1993 to 1998. Self-reported weight, height, lifestyle factors, usual diet, and medical history were collected via an interviewer-administered questionnaire. Incident ESRD cases were identified via record linkage with the nationwide ESRD registry. The computed Cox proportional hazard regression was adjusted for potential risk factors. After an average follow-up of 15.5 years, 827 incident ESRD cases were identified. Compared with a normal BMI of 18.5 to under 23 kg/m2, the hazard ratios and (95% confidence intervals) of ESRD risk for BMIs under 18.5, 23 to under 27.5, and 27.5 kg/m2 or more were 0.54 (0.37-0.79), 1.40 (1.20-1.64) and 2.13 (1.74-2.59), respectively. This significantly trended, linear, dose-dependent association was only present among those with no history of diabetes, hypertension, coronary heart disease, and stroke at baseline, but not significantly among those with any of these co-morbidities. Thus, BMI itself is a risk factor for ESRD in the general population and this association is present in those without pre-existing diabetes, hypertension, coronary heart disease, and stroke.

Brain tumor segmentation based on the U-NET+⁣+ network with efficientnet encoder.

BACKGROUND: Brain tumor is a highly destructive, aggressive, and fatal disease. The presence of brain tumors can disrupt the brain's ability to control body movements, consciousness, sensations, thoughts, speech, and memory. Brain tumors are often accompanied by symptoms like epilepsy, headaches, and sensory loss, leading to varying degrees of cognitive impairment in affected patients. OBJECTIVE: The study goal is to develop an effective method to detect and segment brain tumor with high accurancy. METHODS: This paper proposes a novel U-Net+⁣+ network using EfficientNet as the encoder to segment brain tumors based on MRI images. We adjust the original U-Net+⁣+ model by removing the dense skip connections between sub-networks to simplify computational complexity and improve model efficiency, while the connections of feature maps at the same resolution level are retained to bridge the semantic gap. RESULTS: The proposed segmentation model is trained and tested on Kaggle's LGG brain tumor dataset, which obtains a satisfying performance with a Dice coefficient of 0.9180. CONCLUSION: This paper conducts research on brain tumor segmentation, using the U-Net+⁣+ network with EfficientNet as an encoder to segment brain tumors based on MRI images. We adjust the original U-Net+⁣+ model to simplify calculations and maintains rich semantic spatial features at the same time. Multiple loss functions are compared in this study and their effectiveness are discussed. The experimental results shows the model achieves a high segmention result with Dice coefficient of 0.9180.

[Near-infrared luminescence and energy transfer of ACaPO4 : Eu2+, Nd3+ (A = Li, K, Na)].

Near-infrared (NIR) luminescence phosphors ACaPO4 : Eu2+, Nd2+ (A = Li, K, Na) were prepared by conventional solid state method and the sensitization of Nd3+ near-infrared luminescence by Eu2+ was investigated. The characteristic NIR luminescence of Nd3+ in ACaPO4 matrix is greatly enhanced by co-doping of Eu2+. The fluorescence properties of ACaPO4 : Eu2+, the NIR luminescence properties of ACaPO4 : Eu2+, Nd3+ and the fluorescence lifetime were studied. The effect of emission wavelength of Eu2+ on NIR luminescence of Nd3+ was investigated; The energy transfer mechanism between Eu2+ and Nd3+ was also discussed. Emission peak wavelength of Eu2+ In ACaPO4 matrixes was found red shift with the series of A = Li, K, Na and the extent of the overlap with the different excitation peaks of Nd3+ changes obviously. It was concluded that the emission peak position of Eu2+ is a very important factor for energy transfer, and the optimal wavelength range for Eu2+ --> Nd3+ energy transfer is 500 to 550 nm.

Feasibility and effect of a guided self-disclosure intervention designed to facilitate benefit finding in breast cancer patients: A pilot study.

PURPOSE: This pilot study aimed to evaluate the feasibility and effect of a guided self-disclosure intervention (GSDI) promoting benefit finding (BF) for breast cancer patients. METHODS: A total of 40 women with breast cancer were randomized either to a GSDI group, which included a 6-session face-to-face self-disclosure intervention, or to a control group. The Benefit Finding Scale (BFS) was used to measure BF, the Distress Disclosure Index (DDI) was used to measure self-disclosure, and the Impact of Event Scale-Revised (IES-R) was used to measure cognitive reappraisal. The outcomes were evaluated at baseline and the 3rd and 6th months. RESULTS: The GSDI group had more satisfaction (t = 2.35, P = .02) than the control group and had significant group effects of higher BF (t = 2.214, P = .03) and a lower avoidance of the IES-R (t = -2.353, P = .024) at the 3rd month. There was a significant difference of BF (t = 2.036, P = .049) between the two groups at the 6th month, and other outcomes were not significant (P > .05). Intention-to treat (ITT) analysis showed significant time effects for all outcomes (P < .05); there were slightly significant time × group effects for BF (F = 4.15, P = .052) and disclosure (F = 2.719, P = .090). There were no time × group effects for the other outcomes (all P > .05). CONCLUSION: This study suggests that the GSDI intervention may be feasible in the clinic and might improve BF for breast cancer patients. However, future research needs to further refine the intervention and expand the sample to carry out a full-scale randomized controlled trial.

Family resilience, perceived social support, and individual resilience in cancer couples: Analysis using the actor-partner interdependence mediation model.

PURPOSE: This study aimed to examine the impact of family resilience on the individual resilience of couples during cancer and explore the potential mediating role of perceived social support and the moderating role of sex in this association in cancer patient-spouse dyads. METHOD: The participants were 272 cancer patients and their spouses (N = 544) who completed the Family Resilience Assessment Scale, the Perceived Social Support Scale and the Resilience Scale. We adopted the actor-partner interdependence mediation model to examine whether and how patients' and their spouses' family resilience was associated with their own and their partners' perceived social support and individual resilience. RESULTS: The results indicated that the patients' and their spouses' level of family resilience was positively associated with their own individual resilience directly and indirectly by increasing their own perceived social support. The family resilience of the spouses was associated with an increase in the patients' individual resilience only indirectly by increasing the patients' perceived social support. The spouse-actor effects between family resilience and individual resilience differed significantly by sex. CONCLUSION: Enhancing family resilience and perceived social support within the family can improve individual resilience. The findings regarding the sex differences serve as a rationale for gender-based approaches to improving individual resilience in the family context.

A single nanowire sensor for intracellular glucose detection.

Glucose metabolism plays an important role in cell energy supply, and quantitative detection of the intracellular glucose level is particularly important for understanding many physiological processes. Glucose electrochemical sensors are widely used for blood and extracellular glucose detection. However, intracellular glucose detection cannot be achieved by these sensors owing to their large size and consequent low spatial resolution. Herein, we developed a single nanowire glucose sensor for electrochemical detection of intracellular glucose by depositing Pt nanoparticles (Pt NPs) on a SiC@C nanowire and further immobilizing glucose oxidase (GOD) thereon. Glucose was converted by GOD to an electroactive product H2O2 which was further electro-catalyzed by Pt NPs. The glucose nanowire sensor is endowed with a high sensitivity, high spatial-temporal resolution and enzyme specificity due to its nanoscale size and enzymatic reaction. This allows the real-time monitoring of the intracellular glucose level, and the increase of the intracellular glucose level induced by a novel potential hypoglycemic agent, reinforcing its potential application in lowering the blood glucose level. This work provides a versatile method for the construction of enzyme-modified nanosensors to electrochemically detect intracellular non-electroactive molecules, which is of great benefit for physiological and pathological studies.

Factors Associated with Chronic Kidney Disease and Their Clinical Utility in Primary Care Clinics in a Multi-Ethnic Southeast Asian Population.

BACKGROUND: Chronic kidney disease (CKD) is a major global public health challenge. We investigated determinants of CKD and their clinical utility in an ethnically diverse Southeast Asian population. METHODS: Electronic health records (EHR) of adults ≥40 years who visited any one of 4 government polyclinics in Singapore from January 1, 2012 to -December 31, 2015 were analyzed. CKD was defined as an estimated glomerular filtration rate <60 mL/min/1.73 m2 or 1+ dipstick proteinuria excretion, based on 2 measurements ≥3 months apart. CKD-associated factors and their clinical utility for predicting odds of CKD were investigated using multiple logistic regression analysis. RESULTS: Based on the study criteria, 25.9% (95% CI 25.6-26.2) of the 88,765 eligible study individuals had CKD. The factors (OR and 95% CI) independently associated with CKD were older age ≥65 years (2.54 [2.44-2.64] vs. ≤65 years), respectively; men (1.13 [1.09-1.18]); Malay (1.27 [1.20-1.33]) and Indian (0.77 [0.71-0.83]) vs. Chinese ethnicity; overweight (body mass index [BMI] ≥27.5 kg/m2; 1.10 [1.04-1.16]) vs. normal weight (BMI 18 to <23 kg/m2); government (1.22 [1.15-1.31]) vs. private housing; and with hypertension (3.32 [3.09-3.56]), diabetes (6.93 [6.67-7.20]) or stroke (1.46 [1.36-1.56]) vs. without each co-morbidity, respectively. The area under the receiver operating characteristic curve (95% CI) for the model to predict the probability of CKD using hypertension, diabetes, and age was 0.808 (0.805-0.811). Only 28.5% (27.9-29.1%) of individuals with CKD had physician documentation of their CKD status. However, documentation of CKD status was associated with age ≥65 years (1.11 [1.04-1.20] vs. <65 years), men (1.35 [1.26-1.44]) vs. women, with vs. without hypertension (1.24 [1.07-1.44]), Indian (0.80 [0.69-0.92]) compared to Chinese ethnicity, ever smokers (0.89 [0.81-0.99]) vs. non-smokers, and those with vs. without stroke (0.83 [0.75-0.93]). CONCLUSIONS: CKD prevalence in our Southeast Asian population is high and under-documented even in high-risk patients. Our findings highlight factors associated with CKD, and the predictive value of hypertension, diabetes, and advancing age as EHR-based screening targets for CKD. Our results also suggest that complementary educational efforts will be needed to increase physician detection and optimize the management of CKD, especially in high risk and marginalized groups across all clinics in Singapore, and possibly in the region.

Amplification and overexpression of Aurora-A in esophageal squamous cell carcinoma.

Aurora-A/BTAK/STK15 gene which encodes a centrosome-associated kinase is located on chromosome 20q13.2, a highly amplified region in various human tumors. Recent studies have demonstrated the overexpression and amplification of Aurora-A in many malignant human cancers. The purpose of this study was to investigate the amplification and expression of Aurora-A in esophageal squamous cell carcinoma. Amplification of Aurora-A was determined by fluorescence in situ hybridization in 7 esophageal cancer cell lines and real-time PCR in 29 esophageal cancer samples. We detected Aurora-A expression in 7 esophageal cancer cell lines and 38 esophageal cancers samples by semi-quantitative reverse transcription-PCR and Western blot hybridization. The amplification of Aurora-A was detected in 27 of 29 (93.1%) esophageal cancer samples and 6 of 7 (85.7%) cancer cell lines. Aurora-A was overexpressed in 27 of 38 (71.1%) esophageal cancer samples and all 7 esophageal cancer cell lines. We conclude that Aurora-A is amplified and overexpressed in esophageal squamous cancer.

Knockdown of c-Myc expression by RNAi inhibits MCF-7 breast tumor cells growth in vitro and in vivo.

INTRODUCTION: Breast cancer is the leading cause of cancer death in women worldwide. Elevated expression of c-Myc is a frequent genetic abnormality seen in this malignancy. For a better understanding of its role in maintaining the malignant phenotype, we used RNA interference (RNAi) directed against c-Myc in our study. RNAi provides a new, reliable method to investigate gene function and has the potential for gene therapy. The aim of the study was to examine the anti-tumor effects elicited by a decrease in the protein level of c-Myc by RNAi and its possible mechanism of effects in MCF-7 cells. METHOD: A plasmid-based polymerase III promoter system was used to deliver and express short interfering RNA (siRNA) targeting c-myc to reduce its expression in MCF-7 cells. Western blot analysis was used to measure the protein level of c-Myc. We assessed the effects of c-Myc silencing on tumor growth by a growth curve, by soft agar assay and by nude mice experiments in vivo. Standard fluorescence-activated cell sorter analysis and TdT-mediated dUTP nick end labelling assay were used to determine apoptosis of the cells. RESULTS: Our data showed that plasmids expressing siRNA against c-myc markedly and durably reduced its expression in MCF-7 cells by up to 80%, decreased the growth rate of MCF-7 cells, inhibited colony formation in soft agar and significantly reduced tumor growth in nude mice. We also found that depletion of c-Myc in this manner promoted apoptosis of MCF-7 cells upon serum withdrawal. CONCLUSION: c-Myc has a pivotal function in the development of breast cancer. Our data show that decreasing the c-Myc protein level in MCF-7 cells by RNAi could significantly inhibit tumor growth both in vitro and in vivo, and imply the therapeutic potential of RNAi on the treatment of breast cancer by targeting overexpression oncogenes such as c-myc, and c-myc might be a potential therapeutic target for human breast cancer.

[Expression of survivin and its significance in esophageal cancer].

OBJECTIVE: To detect the expression of survivin in esophageal cancer and elucidate its function in esophageal cancer. METHODS: Expression of surviv in was detected in paired normal and tumor tissues from patients with esophageal cancer by semi-quantitative RT-PCR. A dominant-negative survivin (surT34A) was transfected into esophageal cancer EC9706 cells (EC9706surT34A). Colony formation and apoptosis of the parental and surT34A-transfected EC9706 cells were examined in soft agar and by flow cytometry, respectively. RESULTS: Survivin mRNA expression of tumor tissues was higher than normal tissues in 18/27 (66.7%) samples. The expression level of survivin mRNA in tumor tissues (2.08 +/- 1.32) was significantly higher than that in normal tissues (1.22 +/- 1.09). EC9706 surT34A cells formed fewer colonies on agar than the non-transfected ones. After serum withdrawal, EC9706surT34A had higher apoptotic ratio than control, but survivin could reduce the apoptotic ratio. CONCLUSION: Overexpression of survivin is a common eventin esophageal cancer. The dominant-negative survivin can partially inhibit the malignant phenotype of esophageal cancer.

Detection of human papillomavirus in Chinese esophageal squamous cell carcinoma and its adjacent normal epithelium.

AIM: To investigate the putative role of human papillomavirus (HPV) infection in the carcinogenesis of esophageal squamous cell carcinoma in China. METHODS: Twenty-three esophageal squamous cell carcinoma samples and the distal normal epithelium from Shanxi Province, and 25 more esophageal squamous cell carcinoma samples from Anyang city, two areas with a high incidence of esophageal cancer in China, were detected for the existence of HPV-16 DNA by PCR, mRNA in situ hybridization (ISH) and immunohistochemistry (IHC) targeting HPV-16 E6 gene. RESULTS: There were approximately 64 % (31/48) patients having HPV-16 DNA in tumor samples, among them nearly two-thirds (19/31) samples were detected with mRNA expression of HPV-16 E6. However, in the normal esophageal epithelium from cancer patients, the DNA and mRNA of HPV-16 were found with much less rate: 34.7 % (8/23) and 26.1 % (6/23) respectively. In addition, at protein level detected by IHC assay, 27.1 % (13/48) tumor samples had virus oncoprotein E6 expression, while only one case of normal epithelium was found positive. CONCLUSION: HPV infection, especially type 16, should be considered as a risk factor for esophageal malignancies in China.