Multifunctional fluorescence probe for simultaneous detection of viscosity, polarity, and ONOO- and its bioimaging in vitro and vivo.

Intracellular microenvironment (viscosity and polarity) and peroxynitrite ions (ONOO-) are involved in maintaining cell morphology, cell function, and signaling so that it is crucial to explore their level changes in vitro and vivo. In this work, we designed and synthesized a mitochondria-targeted fluorescence probe XBL for monitoring the dynamic changes of viscosity, polarity, and ONOO- based on TICT and ICT mechanism. The fluorescence spectra showed obvious changes for polarity at 500 nm as well as ONOO- and viscosity at 660 nm, respectively. The XBL can image simultaneously viscosity, polarity, and ONOO- in cells, and the results showed excess ONOO- leaded to the increase of viscosity in mitochondrial. The ferroptosis process was accompanied by increase of intracellular viscosity and ONOO- levels (or decrease of polarity), which allowed us to better understand the relevant physiological and pathological processes. The XBL can distinguish normal cells and cancerous cells by the fluorescence intensity changes in green and red channels, and image viscosity in inflamed mice. Thus, XBL can provided the chemical tool to understand the physiological and pathological mechanisms of disease by simultaneous detection of viscosity, polarity and ONOO-.

A highly selective probe for ratiometric imaging peroxynitrite in living cells and in vivo.

In this study, we constructed and displayed a ratiometric fluorescent probe JQ-2 for detecting ONOO-. The probe JQ-2 showed a ratiometric signal for visualizing ONOO- with a rapid response and high selectivity over a panel of biological analytes. Moreover, the JQ-2 has near-infrared emission (657 nm), which provides an excellent basis for the practical application in biological systems. The probe JQ-2 possessed low cytotoxicity and excellent cell membrane permeability, which can specifically visualize the exogenous and endogenous ONOO- in vitro and vivo by emission in two channels. Meanwhile, JQ-2 can be used for diagnosing drug-induced liver injury by visualizing and monitoring the fluctuations of endogenous ONOO-. Therefore, JQ-2 provided a potential tool for precisely detecting the fluctuation of ONOO- in biological systems to understand physiological and pathological process.

Detection of drug resistance genes of mycobacterium tuberculosis by rolling circle amplification technique with multicolor fluorescent probes / 中华预防医学杂志

The aim of this study was to construct a simple, rapid and ultra-sensitive optical biosensing technique based on rolling circle amplification (RCA), and to apply it to multiple detection of drug-resistant genes of mycobacterium tuberculosis. The common mutation sites of isoniazid, rifampicin and streptomycin resistance genes are katG315 (AGC➝ACC), rpoB531 (CAC➝TAC) and rpsL43 (AAG➝AGG). For these three gene sites, from February 2020 to May 2021, in the Department of Laboratory Medicine of the First Affiliated Hospital of Army Military Medical University, the padlock probe (PLP), primers and capture probes were designed. And a solid-phase RCA constant temperature amplification reaction system based on magnetic beads was constructed and the experimental parameters were optimized. The RCA products were accurately captured by the multicolor fluorescent probes (Cy3/Cy5/ROX), and the single-tube multiple detection of three mutation genes was realized. The sensitivity, specificity and linear range of this method were further verified. The results showed that the response range of katG315 in the same reaction system ranged from 1.0 pmol/L to 0.1 nmol/L. The response range of rpoB531 and rpsL43 ranged from 1.0 pmol/L to 50.0 pmol/L and 1.0 pmol/L to 20.0 pmol/L, and the method had good specificity and sensitivity, and could accurately identify single base mutations in mixed targets, with the minimum detection limit as low as 1.0 pmol/L. The recoveries of simulated serum samples were 95.0%-105.2%. In conclusion, the constant temperature amplification multiple detection method constructed in this study can quickly realize the single-tube multiple detection of three drug resistance mutation sites. This technology is low-cost, simple and rapid, and does not rely on large equipment, providing a new analysis method for pathogen drug resistance gene detection.

Effect of CDK1 Interferes with the Regulation of PLK1, Aurora B and TRF1 on the Proliferation of Leukemia Cells / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of CDK1 interference regulation of PLK1, Aurora B and TRF1 on the proliferation of leukemia cells.@*METHODS@#The human myelogenous leukemia cell line HL-60 was selected as the research object, and the effect of TRF1 expression and its changes on cell proliferation and cycle was investigated by regulating intracellular CDK1 expression. The objects were divided into 5 groups, including control group, shRNA-NC group, CDK1-shRNA group, pcDNA group and pcDNA-CDK1 group. RT-PCR was used to detect the CDK1 expression of cells in each group; colony formation was used to detect the proliferation of the cells. Western blot was used to detect the expression of CDK1, PLK1, Aurora B, TRF1, and cyclin p53, p27, cyclinA.@*RESULTS@#The phosphorylation level of PLK1, Aurora B and the expression of TRF1 in the CDK1-shRNA group were significantly down-regulated as compared with those in the control group (P<0.05). Compared with the control group, the cells in CDK1-shRNA group showed lower clone formation rate, the increasing of cycle-associated proteins p53 and p27 and the decreasing of cyclinA expression (P<0.05). It was shown that interfered CDK1 expression could inhibit the proliferation of HL-60 cells and prolong the time that they enter mitosis, thereby extending the cell cycle. Compared with the control group, the overexpressed CDK1 in the pcDNA-CDK1 group made the phosphorylation level of PLK1, Aurora B, and TRF1 expression increase significantly (P<0.05), also the colony formation rate (P<0.05). The cycle-related proteins p53 and p27 was down-regulated, while cyclinA expression was up-regulate significantly (P<0.05). The results indicted that overexpressed CDK1 could stimulate adverse reactions, thereby promoting the proliferation of HL-60 cells and shortening the cell cycle.@*CONCLUSION@#Knocking out CDK1 can inhibit the phosphorylation of PLK1 and Aurora B and negatively regulate TRF1, thereby inhibiting the proliferation of leukemia cells.

The efficacy and safety of ticagrelor versus clopidogrel in elderly patients with acute coronary syndrome undergoing percutaneous coronary intervention / 中华心血管病杂志

Objective: To compare the efficacy and safety of ticagrelor and clopidogrel in elderly Chinese patients with acute coronary syndrome (ACS) underwent percutaneous coronary intervention (PCI) in the real world. Methods: This study is a post-hoc analysis of a single center, retrospective cohort study. Between March 2016 and March 2018, elderly (age≥65) ACS patients who underwent PCI in the General Hospital of Northern Theater Command were included in the study. The patients were grouped according to P2Y12 receptor inhibitor. The primary endpoints of this study were ischemic events during the 2-year follow-up, which were defined as the composite of cardiac death, myocardial or stroke. The secondary efficiency endpoints included all-cause death and BARC 2, 3, 5 bleeding events. Results: A total of 4 022 elderly (mean age: (71.5±5.3) years) ACS patients were included in this study. Based on the choice of P2Y12 receptor inhibitor, patients were divided into clopidogrel (n=3 201) and ticagrelor (n=821) groups. Incidences of ischemic events (3.2% (26/821) vs. 5.6% (179/3 201), P=0.005) at 2 years were significantly lower in ticagrelor group compared to clopidogrel group. BARC 2, 3, 5 bleeding events (1.7% (14/821) vs. 1.6% (52/3 201), P=0.818) were comparable between the two groups. The incidence of all-cause death (1.5% (12/821) vs. 4.1% (132/3 201), P=0.005) were also lower in the ticagrelor group compared to the clopidogrel group. Clinical outcomes were consistent after adjusting for confounding factors, the incidence of ischemic events (HR= 0.637, 95%CI 0.409-0.991, P=0.046) and all-cause mortality (HR=0.402, 95%CI 0.213-0.758, P=0.005) was significantly lower in the ticagrelor group compared with the clopidogrel group. Risk of BARC 2, 3, 5 bleeding events were similar between the two groups (HR=0.957, 95%CI 0.496-1.848, P=0.897). Conclusion: In real-world clinical practice, for elderly patients with ACS undergoing PCI, ticagrelor use might reduce the incidence of long-term ischemic events and all-cause death without increasing the risk of bleeding.

[Biochar addition improves soil phosphorus availability: A meta-analysis]. / ç”Ÿç‰©ç‚­æé«˜åœŸå£¤ç£·ç´ æœ‰æ•ˆæ€§çš„æ•´åˆåˆ†æž.

Biochar is a potential source for improving soil fertility and crop yield by enhancing phosphorus (P) availability. But the information on quantitative effect of biochar addition on soil P availability is still limited. To address this query, we conducted a meta-analysis with 507 data from 95 eligible literature. The results showed that irrespective of biochar characters (raw material, C:N ratio, pyrolysis temperature, application rate), soil characteristics (texture, pH, organic carbon content), and fertilizer application, biochar addition significantly improved soil available P content by 57.6%. Meanwhile, biochar addition promoted P utilization of crops. The response ratios of plant P concentration to biochar addition were generally lower than those of soil available P. The average response ratio of plant P concentration was 30.6%. The biochars, derived from livestock manure, low-temperature pyrolysis, with lower C:N ratio, alkaline, or higher application rate, were more effective to improve soil available P content and plant P concentration in sandy and loamy soils. For main enzymes involved in P cycle, biochar addition increased activity of alkaline phosphatase (2.8%) but decreased the acid phosphatase activity (17.8%). Overall, biochar addition positively affects soil available and plant P concentration, but has a minute effect on soil phosphatase. The improvement of soil P availability might mainly be ascribed to a great amount of active P fractions in biochar itself.

Finding a resveratrol analogue as potential anticancer agent with apoptosis and cycle arrest.

Resveratrol has been extensively studied as the anti-cancer agent. A variety of resveratrol analogues have been developed with structural modification to improve its bioactivity. In this work, resveratrol analogues, compound 1-4, were designed and synthesized with the Stille-Heck reaction. These results showed compound 1-4 had better anticancer effect than that of parent resveratrol. Especially compound 1 ((E)-4,4'-(ethene-1,2-diyl)bis(3-methylphenol)) displayed the excellent cytotoxicity and high selectivity. The mechanism research indicated compound 1 inhibited cell proliferation by binary paths of cell cycle arrest in S phase regulated by cyclin A1/A2 and apoptosis induction mediated by Bax/Bcl2 in a prooxidant manner.

Finding an efficient tetramethylated hydroxydiethylene of resveratrol analogue for potential anticancer agent.

With the improvement and advance in cancer diagnosis and treatment, the cancer is still a major cause of morbidity and mortality throughout the world. Obviously, new breakthroughs in therapies remain be urgent needed. In this work, we designed and synthesized the compound 1-4, namely resveratrol analogues with methylation of hydroxy distyrene, to further explore its new anti-cancer potential. Encouragingly, compound 1 ((E)-4,4'-(ethene-1,2-diyl)bis(3,5-dimethylphenol)) exhibited cytotoxicity superior to resveratrol in MCF 7 cells. More importantly, the compound 1 showed greater toxicity to tumor cells than that to normal cells, which proved that it could selectively kill tumor cells. The favorable results encouraged us to explore the inhibitory mechanism of compound 1 on MCF 7 cells. The research finding indicated the compound 1 inhibited tumor cell proliferation by both arresting cell cycle in S phase and apoptosis via a prooxidant manner. In addition, the results further verified compound 1 caused cell cycle arrest in S phase and apoptosis by down-regulation of the cycling A1/cycling A2 expression and the rise of Bax/Bcl-2 ratio in a p21-dependant pathway in MCF 7 cells. Therefore, these results are helpful for the effective design of anticancer reagents and the better understanding of their mechanism of action.

REBACIN® as a noninvasive clinical intervention for high-risk human papillomavirus persistent infection.

The development of highly sensitive HPV-genotyping tests has opened the possibility of treating HPV-infected women before high-grade lesions appear. The lack of efficient intervention for persistent high-risk HPV infection necessitates the need for development of novel therapeutic strategy. Here we demonstrate that REBACIN®, a proprietary antiviral biologics, has shown potent efficacy in the clearance of persistent HPV infections. Two independent parallel clinical studies were investigated, which a total of 199 patients were enrolled and randomly divided into a REBACIN®-test group and a control group without treatment. The viral clearance rates for the REBACIN® groups were 61.5% (24/39) and 62.5% (35/56), respectively, for the two independent parallel studies. In contrast, the nontreatment groups showed self-clearance rates at 20.0% (8/40) and 12.5% (8/64). We further found that REBACIN® was able to significantly repress the expression of HPV E6 and E7 oncogenes in TC-1 and Hela cells. The two viral genes are well known for the development of high-grade premalignancy lesion and cervical cancer. In a mouse model, REBACIN® was indicated to notably suppress E6/E7-induced tumor growth, suggesting E6 and E7 oncogenes as a potential target of REBACIN®. Taken together, our studies shed light into the development of a novel noninvasive therapeutic intervention for clearance of persistent HPV infection with significant efficacy.

A near-infrared fluorescent probe for direct and selective detection of cysteine over homocysteine and glutathione.

In this work, we have designed and synthesized the fluorescent probe 1, which showed a highly selective and sensitive response to Cys over Hcy/GSH in the test. Moreover, the color of probe solution has changed dramatically from colorless to pink with the addition of Cys within 10 min. Meanwhile, the fluorescence intensity exhibited perfectly positive correlation with concentration of Cys from 0 to 200 µM, which offered the important condition for quantitative analysis. Finally, the bioimaging and fluorescence response of probe 1 for fetal calf serum are a powerful safeguard for practical detection of Cys. Therefore, this near-infrared probe will be of great benefit for detecting Cys in the biological systems.

[Hyperspectral parameters and prediction model of soil moisture in coastal saline].

Based on the data of soil moisture content and indoor soil surface spectral reflectance from five sampling sites of coastal saline soil, this paper analyzed the relationship between soil moisture content and soil spectrum in wavelength 350-2500 nm. We determined spectral parameters under ratio spectral index (RSI), normalized difference spectral index (NDSI) and difference spectral index (DI), and established the quantitative model of soil moisture content. The results showed significant negative correlation between spectral reflectance and soil moisture content, and the maximum negative correlation was near 1930 nm (r=0.86). By comparison of the regression equation of RSI, NDSI and DI, it was found that the regression equation of exponential function (y=0.00001e9.7203x) built by soil moisture content based on RSI (R1407, R1459) presented the maximum R2 (0.780) and the minimum SE (0.016). The established model based on RSI (R1407, R1459) could be used to monitor soil moisture content accurately in Jiangsu coastal saline soils.

Establishment of reference intervals and transfusion criterion for Sonoclot analysis / 华中科技大学学报（医学）（英德文版）

Sonoclot analyzer has been widely used in many countries. But the reference intervals provided by the manufacturer were derived from only 45 participants, and there was no cut-off value for transfusion for Sonoclot analysis. This study aimed to establish reference intervals and transfusion criterion for Sonoclot analysis. Volunteers were recruited from healthy Chinese adults and patients undergoing cardiac surgery. Blood samples were withdrawn from forearm vein and measured for activated clotting time (ACT), clot rate (CR), platelet function (PF), activated partial thromboplastin time (APTT), fibrinogen concentration (FIB), and platelet count (PLT). The reference intervals were determined by the nonparametric method. Cut-off values were determined by the receiver operating characteristics curve. A total of 135 healthy volunteers and 281 patients were enrolled. The 95% reference intervals were 96-195 s, 22-51 signal U/min, >1.6 for ACT, CR, PF respectively. In the 281 patients, the results of APTT, FIB, PLT, ACT, CR, and PF ranged from 20.5-300.0 s, 0.28-4.11 g/L, (19.0-387.3)×109/L, 80-514 s, 2.9-74 signal U/min, and 0.1-5.1 respectively. The cut-off values for transfusion were >208, ≤14, and ≤1.3 for ACT, CR, PF respectively. The cut-off values of Sonoclot analysis were within the manufacturer's reference intervals, while they were outside the reference intervals established in this study. The results suggested that the manufacturer's reference intervals were not suitable for Chinese. The reference intervals and cut-off values established in this study will be helpful to Chinese patients.

Pathological observation of acute myocardial infarction in Chinese miniswine.

The acute myocardial infarction (AMI) model in Chinese miniswine was built by percutaneous coronary artery occlusion. Pathological observation of AMI was performed, and the expression of tumor necrosis factor alpha (TNF-α) in the infarct sites was detected at different days after modeling in Chinese miniswine. The experimental findings may be used as the basis for blood flow reconstruction and intervention after AMI. Seven experimental Chinese miniswine were subjected to general anesthesia and Seldinger right femoral artery puncture. After coronary angiography, the gelfoam was injected via the microtube to occlude the obtuse marginal branch (OM branch). At 1 d, 3 d, 5 d, 7 d, 10 d, 14 d and 17 d after modeling, hetatoxylin-eosin (HE) staining was performed to observe the pathological changes and to detect the expression of TNF-α in the myocardial tissues. Cytoplasmic acidophilia of the necrotic myocardial tissues at 1 d after modeling was enhanced, and cytoplasmic granules were formed; at 3 d, the margins of the necrotic myocardial tissues were infiltrated by a large number of inflammatory cells; at 5 d, the nuclei of the necrotic myocardial cells were fragmented; at 7 d, extensive granulation tissues were formed at the margin of the necrotic myocardial tissues; at 10 d, part of the granulation tissues were replaced by fibrous scar tissues; at 14-17 d, all granulation tissues were replaced by fibrous scar tissues. Immunohistochemical detection indicated that no TNF-α expression in normal myocardial tissues. The TNF-α expression was first detected at 3 d in the necrotic myocardial tissues and then increased at 5 d and 7 d. After reaching the peak at 10 d, the expression began to decrease at 14 d and the decrease continued at 17 d. Coronary angiography showed the disappearance of blood flow at the distal end of OM branch occluded by gelfoam, indicating that AMI model was constructed successfully. The repair of the infarcted myocardium began at 10-17 d after modeling with safe blood flow reconstruction. TNF-α expression in the infarcted myocardium was the highest at 10 d, which can be explained by inflammation and repair of the infarcted myocardium.

Anti-tumor activity of components isolated from purple sweet potato polysaccharides / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To isolate and purify components from polysaccharides of purple sweet potato (PPSP) and to test their anti-tumor activity.</p><p><b>METHODS</b>DEAE-Cellulose and CM-Cellulose exchange chromatography were applied to separate components of PPSP. The anti-tumor activities of each component were measured by MTT assay on Hela and HepG(2) cells and their monosaccharide composition were analyzed by TLC chromatography, followed by infrared spectroscopy studies.</p><p><b>RESULTS</b>Through weak anion exchange chromatography and gradient elution by sodium chloride solution, four components were separated and named as PPSP, PPSPII, PPSPIII and PPSPIV, respectively. MTT tests showed that PPSP II and PPSPIII inhibited Hela and HepG2 tumor cells in a certain extent. The structural analysis revealed that PPSPI was mainly composed of glucose and galactose, PPSP II was composed of glucose and had a typical absorption peak of β-D-glucose chitosan pyranose, PPSP III was a glycoprotein showing a protein absorption peak.</p><p><b>CONCLUSION</b>Four components were separated from PPSP successfully, among which PPSP II and PPSP III shows anti-tumor activities on Hela and HepG(2) cells in vitro.</p>

PINCH protein expression in normal endometrium, atypical endometrial hyperplasia and endometrioid endometrial carcinoma.

BACKGROUND: Particularly interesting new cysteine-histidine rich protein (PINCH), as an adapter protein of the LIM family for signal transduction in the integrin and growth factor pathway, is upregulated in the stroma of several common types of cancers and involved in promoting tumor progression. In the present study, we examined PINCH expression in normal endometrium, atypical endometrial hyperplasia and endometrioid carcinoma, and further studied the relationships of PINCH expression with clinicopathological variables in cancer patients. METHODS: PINCH expression was examined by immunohistochemistry in 23 normal endometrial samples, 18 atypical endometrial hyperplasias and 48 endometrioid endometrial carcinomas. RESULTS: The PINCH expression in the stroma of cancer (71%) was significantly increased compared to either normal endometrium (17%, p < 0.0001) or atypical hyperplasia (39%, p = 0.017), along with 9 cancers that had stronger PINCH expressions at the invasive margin of the cancers compared to the inner cancers. PINCH expression in cancer was higher in the patients with hypertension (p = 0.041) and estrogen exposure time >30 years (p = 0.021). On the other hand, PINCH expression was not related to menopausal status, gravid status, blood sugar/lipid, family background of cancer, histological grade, myometrial invasion, cervical involvement, lymph nodal metastases, growth pattern, estrogen and progestogen receptors (p > 0.05). conclusion: The results suggest that PINCH seems to play a role, presently unknown, in the tumorigenesis and development of endometrial cancer that merits further study.

[Modeling of cotton boll maturation period and cottonseed biomass accumulation].

Field experiments with different maturity cotton cultivars and sowing dates were conducted at different sites to quantitatively study the effects of cultivar characteristics, weather conditions (air temperature and solar radiation), and crop management variable (N application rate) on the cotton boll maturation period and cottonseed biomass accumulation. The cotton boll maturation period was simulated by using the scale of physiological development time. Based on the hypothesis of sink-determined, the cottonseed biomass accumulation model was then developed. The subtending leaf N concentration of cotton boll was simulated with a semi-empirical equation, and used as the direct indicator of the N nutrition effect on cottonseed growth and development. The model was tested by independent field data obtained in the Yellow River Valley (Xuzhou and Anyang) and the lower reaches of Yangtze River Valley (Huaian) in 2005. The simulated values of boll maturation period showed reasonable agreement with observed values, with a root mean square error (RMSE) of 2.25 days for cultivar DSC-1, of 2.61 days for cultivar KC-1, and of 2.75 days for cultivar AC-33B. The RMSE of cottonseed dry mass prediction was 9.5 mg x seed(-1) for KC-1 and 8.2 mg x seed(-1) for AC-33B, indicating that the model had a good prediction precision.