ObjectiveOur recent study has demonstrated that extracellular acidification promotes lipid accumulation in macrophages via the activation of acid sensing ion channel 1 (ASIC1), but the underlying mechanism remains unclear. This study aims to explore the effect of extracellular acidification on macrophage lipophagy and the underlying mechanism. MethodsRAW264.7 macrophages were incubated with 25 mg/Lox-LDL in a pH 6.5 culture medium for 24 h to build macrophage-derived foam cell models induced by extracellular acidification. Then, RAW264.7 macrophages were cultured in the acidic medium of pH 6.5 with or without PcTx-1 (ASIC1 specific blocker, 10 μg/L) or Nec-1 (RIP1 specific inhibitor, 20 μmol/L) for 24 h, intracellular lipid accumulation was observed by oil red O staining. The expressions of total ASIC1, plasma membrane ASIC1, RIP1, p-RIP1 Ser166, TFEB, p-TFEB Ser142, LC3 and p62 were measured by Western blot. The co-localization of lipids (indicated by Bodipy) with LC3II (autophagosomes) and LAMP1 (lysosomes) was analyzed by a confocal laser scanning microscopy, respectively. Morphological changes of lipophagy in the cells were observed by using transmission electron microscopy. ABCA1-mediated cholesterol efflux was determined by cholesterol fluorescence kits. ResultsCompared with pH 7.4+ox-LDL group, the intracellular lipid accumulation in the pH 6.5+ox-LDL group was significantly increased. Meanwhile, the expressions of plasma membrane ASIC1, p-RIP1 Ser166, p-TFEB Ser142, and p62 proteins were elevated significantly, while LC3II protein level and LC3II/LC3I ratio were decreased. Accordingly, compared with pH 7.4+ox-LDL group, the macrophage lipophagy of the pH 6.5+ox-LDL group was inhibited as indicated by the decreased localization of lipid droplets with LC3 and LAMP1, a decrease in the number of lipophagosomes as well as an increase in lipid droplets. Furthermore, ATP binding cassette transporter A1 (ABCA1)-dependent cholesterol efflux from the macrophages of pH 6.5+ox-LDL group reduced dramatically. However, these above effects of extracellular acidification on RAW264.7 macrophages were abolished by PcTx-1 and Nec-1, respectively. ConclusionThese findings suggest extracellular acidification promotes the phosphorylation of TFEB at Ser142 via activating ASIC1/RIP1 pathway, thereby impeding lipophagy in RAW 264.7 macrophages, and that ASIC1 may be a new potential target for preventing aberrant lipid accumulation diseases including atherosclerosis.

Magnetic beads (MB),composed of magnetic nanoparticles and biocompatible materials including cellulose,chitosan,silica and polystyrene,are ideal platforms for specific detection of cancer cells,virus and nucleic acids when loaded with antibodies,fluorescent and conjugation molecules. With the ease of separations using magnets,magnetic particles show tremendous advantages over other platforms and thus arising potentials in circulating tumor cell (CTC),virus (such as SARS-CoV-2) antibody and antigen detections,and nucleic acid extractions and purifications. In addition,when combined with microfluidic devices,magnetic particles can be used to develop detection platforms with high throughputs,as well as portable and point-of-care apparatus. In accompany with their huge potentials in all kinds of biomedical and clinical scenarios,magnetic particles based medical devices have been commercialized such as antigen test kits that have been commonly used during the pandemic. In this review,the progress in detailed antibody conjugation practice used in each application scenario based on MB platform was summarized and the commercialization of analytical devices combined with microfluidics and others was discussed.

A 31P-quantitative nuclear magnetic resonance(qNMR)method was established for simultaneous determination of sodium α-glycerophosphate,sodium β-glycerophosphate and phosphoric acid in concentrated divitamins and sodium phosphate syrup.The qNMR experimental conditions were optimized,including hexamethylphosphoramide as internal standard,20%deuterium oxide solution as solvent,zgig pulse sequence,delay time of 30 s,and scan number of 64.The 31P-NMR peaks atδ29.80 of hexamethylphosphoramide,δ0.69 of sodiumα-glycerophosphate,δ0.17 of sodiumβ-glycerophosphate andδ0.03 of phosphoric acid were chosen as the quantitative peaks(pH of the test solution was around 4.8).Method validation was performed in terms of precision(Relative standard deviation less than 0.6%),linearity(Correlative coefficient greater than 0.999),limit of detection(23.58 μg/mL for sodium glycerophosphate and 11.61 μg/mL for phosphoric acid)and limit of quantitation(78.60 μg/mL for sodium glycerophosphate and 38.70 μg/mL for phosphoric acid).The recoveries were 99.8%?103.2%,and relative standard deviations were 0.41%?1.98%.The results showed that the reliability of 31P-qNMR method were suitable for its intended use.Seven batches of concentrated divitamins and sodium phosphate syrups were tested by the established method,of which the total phosphorus content was consistent with that of colorimetry method,but the content of sodium glycerophosphate(Sum ofαtype andβtype)was relatively low,about 82%of the labeled amount.The content of phosphoric acid was high.This method simplified sample pretreatment and had high specificity,and was more suitable for determination and quality control of concentrated divitamins and sodium phosphate syrup.

AIM To observe the effects of Yiqi Fumai Lyophilized Injection on a mouse model of chronic heart failure after acute myocardial infarction and H9C2 cells with injury induced by oxygen and glucose deprivation.METHODS Thirty C57BL/6J mice were randomly divided into the sham-operation group,the model group,and the Yiqi Fumai group,among which the chronic heart failure model was made by ligation of anterior descending coronary artery and given 4 weeks corresponding drug administration.The H9C2 cells were divided into the normal group,the OGD model group,the low-dose,medium-dose and high-dose Yiqi Fumai groups,and the JQ1(BRD4 inhibitor)group.The following detections were conducted:heart ultrasound among the mice;the HE staining of the myocardial tissue for the observation of the morphological changes,spectrophotometry of cellular ATP level,JC-1 method for the determination of the mitochondrial membrane potential,RT-qPCR for the assessment of the mRNA expressions of ANP、BNP and BRD4,TUNEL method for the determination of level of cardiomyocytes apoptosis,and Western blot for the determination of the protein expressions of BRD4,AMPK and p-AMPK in both the mice and H9C2 cells.RESULTS Compared with the sham-operation group,the model group displayed decreased heart function(P＜0.01);obviously more hypertrophic or morphologically irregular cardiomyocytes;up-regulated mRNA expressions of ANP,BNP and BRD4(P＜0.01);increased cardiomyocyte apoptosis rate(P＜0.01);decreased p-AMPK/AMPK protein expression(P＜0.01);and increased BRD4 protein expression(P＜0.01).Compared with the model group,the Yiqi Fumai groups shared improvement in terms of all the aforementioned indices levels(P＜0.05,P＜0.01).Compared with the H9C2 cells of the normal group,those of the OGD model group displayed up-regulated mRNA expressions of ANP,BNP and BRD4(P＜0.01);increased apoptosis rate(P＜0.01);decreased p-AMPK/AMPK protein expression(P＜0.01);increased BRD4 protein expression(P＜0.01),and decreased mitochondrial membrane potential and ATP levels(P＜0.01).Compared with the OGD model group,the groups treated with either Yiqi Fumai or JQ1 demonstrated improvement in terms of the aforementioned indices levels(P＜0.05,P＜0.01).CONCLUSION Yiqi Fumai Lyophilized Injection exhibits its efficacy in improving cardiac function,myocardial energy metabolism,intracellular mitochondria dysfunction and biogenesis in a mouse model of chronic heart failure after myocardial infarction.The effects may be mediated by regulation of BRD4/AMPK signaling pathway.

Objective To analyze the genetic subtypes of human immunodeficiency virus (HIV) and the prevalence of pretreatment drug resistance in the newly reported HIV-infected men in Guangxi. Methods The stratified random sampling method was employed to select the newly reported HIV-infected men aged≥50 years old in 14 cities of Guangxi from January to June in 2020.The pol gene of HIV-1 was amplified by nested reverse transcription polymerase chain reaction and then sequenced.The mutation sites associated with drug resistance and the degree of drug resistance were then analyzed. Results A total of 615 HIV-infected men were included in the study.The genetic subtypes of CRF01_AE,CRF07_BC,and CRF08_BC accounted for 57.4% (353/615),17.1% (105/615),and 22.4% (138/615),respectively.The mutations associated with the resistance to nucleoside reverse transcriptase inhibitors (NRTI),non-nucleoside reverse transcriptase inhibitors (NNRTI),and protease inhibitors occurred in 8 (1.3%),18 (2.9%),and 0 patients,respectively.M184V (0.7%) and K103N (1.8%) were the mutations with the highest occurrence rates for the resistance to NRTIs and NNRTIs,respectively.Twenty-two (3.6%) patients were resistant to at least one type of inhibitors.Specifically,4 (0.7%),14 (2.3%),4 (0.7%),and 0 patients were resistant to NRTIs,NNRTIs,both NRTIs and NNRTIs,and protease inhibitors,respectively.The pretreatment resistance to NNRTIs had much higher frequency than that to NRTIs (2.9% vs.1.3%;χ²=3.929,P=0.047).The prevalence of pretreatment resistance to lamivudine,zidovudine,tenofovir,abacavir,rilpivirine,efavirenz,nevirapine,and lopinavir/ritonavir was 0.8%, 0.3%, 0.7%, 1.0%, 1.3%, 2.8%, 2.9%, and 0, respectively. Conclusions CRF01_AE,CRF07_BC,and CRF08_BC are the three major strains of HIV-infected men≥50 years old newly reported in Guangxi,2020,and the pretreatment drug resistance demonstrates low prevalence.
Male ; Humans ; Middle Aged ; Reverse Transcriptase Inhibitors/therapeutic use* ; HIV Infections/drug therapy* ; Drug Resistance, Viral/genetics* ; China/epidemiology* ; Mutation ; HIV-1/genetics* ; Protease Inhibitors/therapeutic use* ; Genotype

Both anti-glomerular basement membrane (GBM) disease and the anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) are common causes of pulmonary-renal syndrome. Organizing pneumonia (OP), a special pattern of interstitial lung disease, is extremely rare either in AAV or anti-GBM disease. We report an old woman presented with OP on a background of co-presentation with both ANCA and anti-GBM antibodies.
Female ; Humans ; Antibodies, Antineutrophil Cytoplasmic ; Organizing Pneumonia ; Autoantibodies ; Glomerulonephritis ; Anti-Glomerular Basement Membrane Disease ; Pneumonia ; Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis/complications*

Objective To investigate the titer stability of the harvest solution of severe acute respiratory syndrome coronavirus2(SARS-CoV-2)at 2 ～ 8 ℃ and the inactivation effect of β-propiolactone inactivator on the virus.Methods Three batches of SARS-CoV-2 harvest solution(batch numbers:202111001,202111002 and 202111003)were stored at 2 ～ 8 ℃ for 12 d and sampled every 3 d(0,3,6,9 and 12 d)for detection of the titers by Karber method;Three batches of virus harvest solution equilibrated overnight at 2 ～ 8 ℃ were inactivated by adding β-propiolactone at a volume fraction of 1∶4 000 and detected for the titers at different inactivation time points(0,0.5,1,1.5,2,3,4,8,16 and 24 h),of which samples inactivated for 8,16 and 24 h were taken for inactivation verification,and samples inactivated for 24 h were observed by transmission electron microscope.Results The titers of SARS-CoV-2 decreased with the prolongation of storage time at 2 ～8 ℃,which showed no obvious decrease during 0 ～ 3 d,while decreased from the initial 7.75,6 and 7.5 lgCCID_(50)/mL to5.75,4.625 and 6.25 lgCCID_(50)/mL on day 12,indicating that the virus activity showed a gradual decrease trend at 2 ～8 ℃；With the inactivation time,the virus titer decreased continuously and could not be detected after inactivation for 3 h.Transmission electron microscope observation showed that the inactivated virus particles were intact and the spike protein was evenly distributed.Conclusion The virulence of SARS-CoV-2 stored at 2 ～ 8 ℃ was unstable,so the subsequent inactivation and purification process should be carried out as soon as possible;The titer of virus could not be detected after3 h of inactivation,which provided a reference for the determination of the inactivation process.

Currently,the research or publications related to the clinical comprehensive evaluation of Chinese patent medicine are increasing,which attracts the broad attention of all circles. According to the completed clinical evaluation report on Chinese patent medicine,there are still practical problems and technical difficulties such as unclear responsibility of the evaluation organization,unclear evaluation subject,miscellaneous evaluation objects,and incomplete and nonstandard evaluation process. In terms of evaluation standards and specifications,there are different types of specifications or guidelines with different emphases issued by different academic groups or relevant institutions. The professional guideline is required to guide the standardized and efficient clinical comprehensive evaluation of Chinese patent medicine and further improve the authority and quality of evaluation. In combination with the characteristics of Chinese patent medicine and the latest research achievement at home and abroad,the detailed specifications were formulated from six aspects including design,theme selection,content and index,outcome,application and appraisal,and quality control. The guideline was developed based on the guideline development requirements of China Assoication of Chinese medicine. After several rounds of expert consensus and public consultation,the current version of the guideline has been developed.
Medicine, Chinese Traditional ; Nonprescription Drugs ; Consensus ; China ; Reference Standards ; Drugs, Chinese Herbal

BACKGROUND: The benefits of healthy lifestyles are well recognized. However, the extent to which improving unhealthy lifestyles reduces cardiovascular disease (CVD) risk needs to be discussed. We evaluated the impact of lifestyle improvement on CVD incidence using data from the China-PAR project (Prediction for Atherosclerotic Cardiovascular Disease Risk in China). METHODS: A total of 12,588 participants free of CVD were followed up for three visits after the baseline examination. Changes in four lifestyle factors (LFs) (smoking, diet, physical activity, and alcohol consumption) were assessed through questionnaires from the baseline to the first follow-up visit. Cox proportional hazard models were used to estimate hazard ratios (HRs) and corresponding 95% confidence intervals (CIs). The risk advancement periods (RAPs: the age difference between exposed and unexposed participants reaching the same incident CVD risk) and population-attributable risk percentage (PAR%) were also calculated. RESULTS: A total of 909 incident CVD cases occurred over a median follow-up of 11.14 years. Compared with maintaining 0-1 healthy LFs, maintaining 3-4 healthy LFs was associated with a 40% risk reduction of incident CVD (HR = 0.60, 95% CI: 0.45-0.79) and delayed CVD risk by 6.31 years (RAP: -6.31 [-9.92, -2.70] years). The PAR% of maintaining 3-4 unhealthy LFs was 22.0% compared to maintaining 0-1 unhealthy LFs. Besides, compared with maintaining two healthy LFs, improving healthy LFs from 2 to 3-4 was associated with a 23% lower risk of CVD (HR = 0.77, 95% CI: 0.60-0.98). CONCLUSIONS Long-term sustenance of healthy lifestyles or improving unhealthy lifestyles can reduce and delay CVD risk.

Objective: To establish a nomogram prognostic model for predicting the 5-, 10-, and 15-year overall survival (OS) of non-metastatic renal cell carcinoma patients managed with radical nephrectomy (RN), compare the modelled results with the results of pure pathologic staging, the Karakiewicz nomogram and the Mayo Clinic Stage, Size, Grade, and Necrosis (SSIGN) score commonly used in foreign countries, and stratify the patients into different prognostic risk subgroups. Methods: A total of 1 246 non-metastatic renal cell carcinoma patients managed with RN in Sun Yat-sen University Cancer Center (SYSUCC) from 1999 to 2020 were retrospectively analyzed. Multivariate Cox regression analysis was used to screen the variables that influence the prognosis for nomogram establishment, and the bootstrap random sampling was used for internal validation. The time-receiver operating characteristic curve (ROC), the calibration curve and the clinical decision curve analysis (DCA) were applied to evaluate the nomogram. The prediction efficacy of the nomogram and that of the pure pathologic staging, the Karakiewicz nomogram and the SSIGN score was compared through the area under the curve (AUC). Finally, patients were stratified into different risk subgroups according to our nomogram scores. Results: A total of 1 246 patients managed with RN were enrolled in this study. Multivariate Cox regression analysis showed that age, smoking history, pathological nuclear grade, sarcomatoid differentiation, tumor necrosis and pathological T and N stages were independent prognostic factors for RN patients (all P<0.05). A nomogram model named SYSUCC based on these factors was built to predict the 5-, 10-, and 15-year survival rate of the participating patients. In the bootstrap random sampling with 1 000 iterations, all these factors occurred for more than 800 times as independent predictors. The Harrell's concordance index (C-index) of SYSUCC was higher compared with pure pathological staging [0.770 (95% CI: 0.716-0.823) vs 0.674 (95% CI: 0.621-0.728)]. The calibration curve showed that the survival rate as predicted by the SYSUCC model simulated the actual rate, while the clinical DCA showed that the SYSUCC nomogram has a benefit in certain probability ranges. In the ROC analysis that included 857 patients with detailed pathological nuclear stages, the nomogram had a larger AUC (5-/10-year AUC: 0.823/0.804) and better discriminating ability than pure pathological staging (5-/10-year AUC: 0.701/0.658), Karakiewicz nomogram (5-/10-year AUC: 0.772/0.734) and SSIGN score (5-/10-year AUC: 0.792/0.750) in predicting the 5-/10-year OS of RN patients (all P<0.05). In addition, the AUC of the SYSUCC nomogram for predicting the 15-year OS (0.820) was larger than that of the SSIGN score (0.709), and there was no statistical difference (P<0.05) between the SYSUCC nomogram, pure pathological staging (0.773) and the Karakiewicz nomogram (0.826). The calibration curve was close to the standard curve, which indicated that the model has good predictive performance. Finally, patients were stratified into low-, intermediate-, and high-risk subgroups (738, 379 and 129, respectively) according to the SYSUCC nomogram scores, among whom patients in intermediate- and high-risk subgroups had a worse OS than patients in the low-risk subgroup (intermediate-risk group vs. low-risk group: HR=4.33, 95% CI: 3.22-5.81, P<0.001; high-risk group vs low-risk group: HR=11.95, 95% CI: 8.29-17.24, P<0.001), and the high-risk subgroup had a worse OS than the intermediate-risk group (HR=2.63, 95% CI: 1.88-3.68, P<0.001). Conclusions: Age, smoking history, pathological nuclear grade, sarcomatoid differentiation, tumor necrosis and pathological stage were independent prognostic factors for non-metastasis renal cell carcinoma patients after RN. The SYSUCC nomogram based on these independent prognostic factors can better predict the 5-, 10-, and 15-year OS than pure pathological staging, the Karakiewicz nomogram and the SSIGN score of patients after RN. In addition, the SYSUCC nomogram has good discrimination, agreement, risk stratification and clinical application potential.
Humans ; Nomograms ; Retrospective Studies ; Carcinoma, Renal Cell/pathology* ; Prognosis ; Risk Factors ; Nephrectomy ; Kidney Neoplasms/pathology* ; Necrosis