Twin Nanopipettes for Real-Time Electrochemical Monitoring of Cytoplasmic Microviscosity at a Single-Cell Level.

Cytoplasmic microviscosity (CPMV) plays essential roles in governing the diffusion-mediated cellular processes and has been recognized as a reliable indicator of the cellular response of many diseases and malfunctions. Current CPMV studies are exclusively established by probe-assisted optical methods, which nevertheless necessitate the complicated synthesis and delivery of optical probes into cells and thus the issues of biocompatibility and bio-orthogonality. Using twin nanopipettes integrated with a patch-clamp system, a practical electrochemical single-cell measurement is presented, which is capable of real-time and long-term CPMV detection without cell disruption. Specifically, upon the operation of the twin nanopipettes, the cellular CPMV status, which is correlated to cytoplasmic ionic mobility, could be sensibly transduced via the ionic current passing through the nanosystem. The average CPMV value of HeLa cells was detected as ca. 86 cP. Notably, the correlation between chemotherapy and CPMV alterations makes this approach possible for the real-time and long-term assessment of the evolution of external stimuli, as exemplified by the two natural products taxol and colchicine. Integrated with the patch-clamp setup, this study features the first use of twin nanopipettes for electrochemical CPMV monitoring of single living cells, and it is expected to inspire more interest in the exploitation of dual- and multiple nanopipettes for advanced single-cell analysis.

Nanoporous Semiconductor Electrode Captures the Quantum Dots: Toward Ultrasensitive Signal-On Liposomal Photoelectrochemical Immunoassay.

Liposomal photoelectrochemical (PEC) bioanalysis has recently emerged and exhibited great potential in sensitive biomolecular detection. Exploration of the facile and efficient route for advanced liposomal PEC bioanalysis is highly appealing. In this work, we report the split-type liposomal PEC immunoassay system consisting of sandwich immunorecognition, CdS quantum dots (QDs)-loaded liposomes (QDLL), and the release and subsequent capture of the QDs by a separated TiO2 nanotubes (NTs) electrode. The system elegantly operated upon the protein binding and lysis treatment of CdS QDLL labels within the 96-well plate, and then the CdS QDs-enabled sensitization of TiO2 NTs electrode. Exemplified by cardiac markers troponin I (cTnI) as target, the proposed system achieved efficient activation of TiO2 NTs electrode and thus the signal generation toward the split-type PEC immunoassay. This work features the first use of QDs for liposomal PEC bioanalysis and is expected to inspire more interests in the design and implementation of numerous QDs-involved liposomal PEC bioanalysis.

Ferroelectric Perovskite Oxide@TiO2 Nanorod Heterostructures: Preparation, Characterization, and Application as a Platform for Photoelectrochemical Bioanalysis.

This work reports the first synthesis and characterization of a ferroelectric perovskite oxide-based heterostructure as well as its application for photoelectrochemical (PEC) bioanalytical purposes. Specifically, exemplified by [KNbO3]1- x[BaNi1/2Nb1/2O3-δ] x (KBNNO), the ferroelectric perovskite oxides were prepared by solid-state synthesis, while the TiO2 nanorod (NR) arrays were obtained via a hydrothermal method. Using the technique of pulsed laser deposition (PLD), KBNNO were then deposited on TiO2 NRs to form KBNNO@TiO2 NR heterostructures. Various characterization techniques were applied to reveal compositional and structural information on the as-fabricated sample, and favorable alignment existed between the two components as displayed by the PEC test. In the detection of l-cysteine, the as-fabricated KBNNO@TiO2 NRs demonstrated good performance in terms of sensitivity and selectivity. This work revealed the potential of ferroelectric perovskite oxide and its heterostructures for innovative PEC bioanalytical applications, and we hope it will generate more interest in the development of various ferroelectrics-based heterostructures for advanced PEC bioanalysis.

Berberine Alleviates Ischemic Brain Injury by Enhancing Autophagic Flux via Facilitation of TFEB Nuclear Translocation.

Berberine has been demonstrated to alleviate cerebral ischemia/reperfusion injury, but its neuroprotective mechanism has yet to be understood. Studies have indicated that ischemic neuronal damage was frequently driven by autophagic/lysosomal dysfunction, which could be restored by boosting transcription factor EB (TFEB) nuclear translocation. Therefore, this study investigated the pharmacological effects of berberine on TFEB-regulated autophagic/lysosomal signaling in neurons after cerebral stroke. A rat model of ischemic stroke and a neuronal ischemia model in HT22 cells were prepared using middle cerebral artery occlusion (MCAO) and oxygen-glucose deprivation (OGD), respectively. Berberine was pre-administered at a dose of 100[Formula: see text]mg/kg/d for three days in rats and 90[Formula: see text][Formula: see text]M in HT22 neurons for 12[Formula: see text]h. 24[Formula: see text]h after MCAO and 2[Formula: see text]h after OGD, the penumbral tissues and OGD neurons were obtained to detect nuclear and cytoplasmic TFEB, and the key proteins in the autophagic/lysosomal pathway were examined using western blot and immunofluorescence, respectively. Meanwhile, neuron survival, infarct volume, and neurological deficits were assessed to evaluate the therapeutic efficacy. The results showed that berberine prominently facilitated TFEB nuclear translocation, as indicated by increased nuclear expression in penumbral neurons as well as in OGD HT22 cells. Consequently, both autophagic activity and lysosomal capacity were simultaneously augmented to alleviate the ischemic injury. However, berberine-conferred neuroprotection could be greatly counteracted by lysosomal inhibitor Bafilomycin A1 (Baf-A1). Meanwhile, autophagy inhibitor 3-Methyladenine (3-MA) also slightly neutralized the pharmacological effect of berberine on ameliorating autophagic/lysosomal dysfunction. Our study suggests that berberine-induced neuroprotection against ischemic stroke is elicited by enhancing autophagic flux via facilitation of TFEB nuclear translocation in neurons.

Development of high-efficiency superparamagnetic drug delivery system with MPI imaging capability.

In this study, a high-efficiency superparamagnetic drug delivery system was developed for preclinical treatment of bladder cancer in small animals. Two types of nanoparticles with magnetic particle imaging (MPI) capability, i.e., single- and multi-core superparamagnetic iron oxide nanoparticles (SPIONs), were selected and coupled with bladder anti-tumor drugs by a covalent coupling scheme. Owing to the minimal particle size, magnetic field strengths of 270 mT with a gradient of 3.2 T/m and 260 mT with a gradient of 3.7 T/m were found to be necessary to reach an average velocity of 2 mm/s for single- and multi-core SPIONs, respectively. To achieve this, a method of constructing an in vitro magnetic field for drug delivery was developed based on hollow multi-coils arranged coaxially in close rows, and magnetic field simulation was used to study the laws of the influence of the coil structure and parameters on the magnetic field. Using this method, a magnetic drug delivery system of single-core SPIONs was developed for rabbit bladder therapy. The delivery system consisted of three coaxially and equidistantly arranged coils with an inner diameter of Φ50 mm, radial height of 85 mm, and width of 15 mm that were positioned in close proximity to each other. CCK8 experimental results showed that the three types of drug-coupled SPION killed tumor cells effectively. By adjusting the axial and radial positions of the rabbit bladder within the inner hole of the delivery coil structure, the magnetic drugs injected could undergo two-dimensional delivery motions and were delivered and aggregated to the specified target location within 12 s, with an aggregation range of about 5 mm × 5 mm. In addition, the SPION distribution before and after delivery was imaged using a home-made open-bore MPI system that could realistically reflect the physical state. This study contributes to the development of local, rapid, and precise drug delivery and the visualization of this process during cancer therapy, and further research on MPI/delivery synchronization technology is planned for the future.

[Inhibitory effect of cocoomyxa gloeobotrydifomis on benign prostate hyperplasia in aged rats and its action mechanism].

OBJECTIVE: To investigate the inhibitory effect of cocoomyxa gloeobotrydifomis (CGD) on benign prostate hyperplasia (BPH) in aged rats and its underlying mechanism. METHODS: Thirty SD male rats aged 21 months were equally randomized to three groups, aged control, low-dose CGD and high-dose CGD, the latter two groups fed on a diet with CGD at 50 and 100 mg per kg per d for 3 months, while the aged controls on normal laboratory chow. Another 10 3-month-old male rats were included in a young control group and fed on the same diet as the aged control rats. At the end of 3 months of CGD treatment, the prostates of all the rats were harvested and weighed. The histomorphological and interstitial changes of the prostatic tissue were observed by HE staining and Masson staining, respectively. The expressions of phosphorylated phosphoinositide-dependent kinase 1 (PDK1), phosphorylated Akt (Ser 473) and phosphorylated PTEN in the rat prostate were determined by Western blotting. RESULTS: The wet weight and index of the prostate were significantly higher in the aged controls than in the young controls ([1 220 +/- 140] vs [550 +/- 60] mg, P < 0.01; 2.08 +/- 0.17 vs 1.94 +/- 0.10, P < 0.05). High-dose CGD significantly inhibited the increase in the prostatic wet weight and index of the aged rats ([1 080 +/- 97] mg and 1.85 +/- 0.16) as compared with the aged controls (P < 0.01 and P < 0.05). The epithelium and interstitium, particularly the latter, were evidently thicker in the aged control than in the CGD-treated rats. The protein levels of phosphorylated PDK1 and Akt were significantly enhanced, while that of phosphorylated PTEN remarkably down-regulated in the aged rats as compared with the young ones. The expressions of phosphorylated PDK1 and Akt were significantly decreased, whereas that of phosphorylated PTEN markedly increased in both the low-dose and high-dose CGD groups. CONCLUSION: CGD can significantly inhibit BPH in aged rats through down-regulating the PI3K/Akt pathway.

A Fast and Reversible Responsive Bionic Transmembrane Nanochannel for Dynamic Single-Cell Quantification of Glutathione.

Biological channels can rapidly and continuously modulate ion transport behaviors in response to external stimuli, which play essential roles in manipulating physiological and pathological processes in cells. Here, to mimic the biological channels, a bionic nanochannel is developed by synergizing a cationic silicon-substituted rhodamine (SiRh) with a glass nanopipette for transmembrane single-cell quantification. Taking the fast and reversible nucleophilic addition reaction between glutathione (GSH) and SiRh, the bionic nanochannel shows a fast and reversible response to GSH, with its inner-surface charges changing between positive and negative charges, leading to a distinct and reversible switch in ionic current rectification (ICR). With the bionic nanochannel, spatiotemporal-resolved operation is performed to quantify endogenous GSH in a single cell, allowing for monitoring of intracellular GSH fluctuation in tumor cells upon photodynamic therapy and ferroptosis. Our results demonstrate that it is a feasible tool for in situ quantification of the endogenous GSH in single cells, which may be adapted to addressing other endogenous biomolecules in single cells by usage of other stimuli-responsive probes.

Organic Molecular Probe Enabled Ionic Current Rectification toward Subcellular Detection of Glutathione with High Selectivity, Sensitivity, and Recyclability.

Single-cell interrogation with the solid-state nanoprobes enables understanding of the linkage between cellular behavior and heterogeneity. Herein, inspired by the charge property of the organic molecular probe (OMP), a generic ionic current rectification (ICR) single-cell methodology is established, exemplified by subcellular detection of glutathione (GSH) with high selectivity, sensitivity, and recyclability. The as-developed nanosensor can transduce the subcellular OMP-GSH interaction via a sensitive ionic response, which stems from the superior specificity of OMP and its essential charge property. In addition, the nanosensor exhibits good reversibility, since the subsequent tandem reaction after the recognition can well recover the sensing surface. Given the diverse structures and tailorable charge properties of OMP, this work underpins a new and general method of OMP-based ICR single-cell analysis.

[Greenlight photoselective vaporization prostatectomy versus thulium laser vaporesection of the prostate for aged high-risk BPH patients with the prostate heavier than 80 g].

OBJECTIVE: To compare the effects of greenlight photoselective vaporization prostatectomy (PVP) and thulium laser vaporesection of the prostate (TmLRP) in the treatment of aged high-risk BPH patients with the prostate weighing > 80 g. METHODS: We included in this study 118 high-risk BPH patients aged 62-96 (mean 76) years with the prostate heavier than 80 g, 82 treated by PVP and the other 36 by TmLRP. Then we compared the operation time, intraoperative bleeding, complications, short-term effectiveness, and surgical cost between the two groups. RESULTS: All the patients tided over the perioperative period without blood transfusion and serious complications. The mean operation time, postoperative bladder irrigation time and surgical cost were significantly less in the TmLRP than in the PVP group (P < 0.05). Both the procedures remarkably improved the international prostatic symptom score (IPSS), quality of life (QOL), post void residual urine (PVR) and Qmax of the patients (P < 0.05), but with no significant differences between the two groups (P > 0.05). CONCLUSION: Both PVP and TmLRP are effective and safe for the treatment of aged high-risk BPH patients with the prostate heavier than 80 g, but the latter is superior for its shorter operation time and lower surgical cost.

[Transurethral thulium laser urethrotomy for urethral stricture].

OBJECTIVE: To evaluate the effect of endourethrotomy with thulium laser as a minimally invasive treatment for urethral stricture. METHODS: We treated 36 cases of urethral stricture or atresia by endourethrotomy with thulium laser, restored the urethral continuity by vaporization excision of the scar tissue, and observed the clinical effects and complications. RESULTS: The mean operation time was 35 min, ranging from 10 to 90 min. Smooth urination was achieved after 2-6 weeks of catheter indwelling, with no urinary incontinence. The patients were followed up for 4-24 (mean 12) months, during which 27 did not need any reintervention, 5 developed urinary thinning but cured by urethral dilation, 3 received another laser urethrotomy for previous negligence of timely urethral dilation, and the other 1 underwent open urethroplasty. CONCLUSION: Thulium laser urethrotomy is a safe and effective minimally invasive option for short urethral stricture, which is also suitable for severe urethral stricture and urethral atresia. Its short-term outcome is satisfactory, but its long-term effect remains to be further observed.

[Study on the production of alkaloid by cell mass suspension culture of Fritillaria cirrhosa].

OBJECTIVE: Set up Fritillaria cirrhosa cell mass suspension culture system to rapidly screen the best culture conditions for cell mass proliferation and hormone combination. METHODS: Using MS medium as the basic medium, the impact of inoculum size, hormone combination, growth regulators for Fritillaria cirrhosa cell mass suspension culture were compared, and also the growth of cell mass at different culture conditions was compared, and the total alkaloids content in proliferative cell mass was measured. RESULTS AND CONCLUSION: Fritillaria cirrhosa grow significantly faster in cell mass suspension culture than in the solid culture. The total alkaloid content in cell mass is higher than commercial and wild bulb of Fritillaria cirrhosa. The optimal inoculum size for cell mass suspension culture is 30 g/L and the optimal culture media is MS +6-BA 2.0 mg/L + NAA 0.2 mg/L.

LINC00511/miRNA-143-3p Modulates Apoptosis and Malignant Phenotype of Bladder Carcinoma Cells via PCMT1.

Bladder cancer has easy recurrence characteristics, but its occurrence and development mechanism are still unclear. Non-coding RNA is a kind of RNA that exists widely and cannot be translated into proteins, which has played a key role in the regulation of biological functions of tumor cells. However, the regulation mechanism of non-coding RNA on bladder tumors is not fully understood. By microarray analysis and database analysis, we found that LINC00511 was significantly highly expressed in bladder cancer. The expressions of LINC00511, miR-143-3p, and PCMT in bladder cancer tissues and cells were detected by quantitative reverse transcription-polymerase chain reaction. The relationship between the expressions of miR-143-3p and PCMT1 and the clinicopathological parameters of the tumor was analyzed. The proliferation and invasion of bladder cancer cells were detected by MTT assay and Transwell assay. The expression levels of E-cadherin and vimentin in bladder cancer cells were detected by Western blot. Cell apoptosis was detected by flow cytometry. In vivo, TCCSUP or SW780 cells were inoculated into BALB/c nude mice to detect tumor volume and weight. Bioinformatics and dual luciferase reporter gene were used to analyze the relationship between LINC00511 and miR-143-3p and its downstream target gene PCMT1. The results showed that LINC00511 could target miR-143-3p/PCMT1 to regulate the proliferation, migration, and apoptosis of bladder cancer TCCSUP or SW780 cells and promote the occurrence and development of bladder cancer.

Photocontrolled Nanopipette Biosensor for ATP Gradient Electroanalysis of Single Living Cells.

Emerging nanopipette tools have demonstrated substantial potential for advanced single-cell analysis, which plays vital roles from fundamental cellular biology to biomedical diagnostics. Highly recyclable nanopipettes with easy and quick regeneration are of special interest for precise and multiple measurements. However, existing recycle strategies are generally plagued by operational complexity and limited efficiency. Light, acting in a noncontact way, should be the ideal external stimulus to address this issue. Herein, we present the photocontrolled nanopipette capable of probing cellular adenosine triphosphate (ATP) gradient at single-cell level with good sensitivity, selectivity, and reversibility, which stems from the use of ATP-specific azobenzene (Azo)-incorporated DNA aptamer strands (AIDAS) and thereby the sensible transduction of variable nanopore size by the ionic currents passing through the aperture. Photoisomerized conformational change of the AIDAS by alternative UV/vis light stimulation ensures its noninvasive regeneration and repeated detection. Inducement and inhibition of the cellular ATP could also be probed by this nanosensor.

Coronary microcirculation changes during myocardial stunning in dogs.

OBJECTIVE: To investigate changes in the coronary microcirculation during myocardial stunning in dogs. METHODS: Male mongrel dogs underwent a 15- or 60-min occlusion of the left anterior descending coronary artery, followed by a 120-min reperfusion. Myocardial contrast echocardiography was performed before and after treatment with acetylcholine (ACH) or nitroglycerin (NG). Peak videointensity (PVI) in the myocardial zone was measured, and myocardial samples were examined using transmission electron microscopy. RESULTS: In the 15-min group, the ratio of the PVI between the stunned and intact myocardial zone (PVIR) before and after treatment with NG (NG-PVIR) or ACH (ACH-PVIR) declined markedly in the early period of reperfusion and then returned to preligation levels. In the 60-min group, NG-PVIR was reduced in the early period of reperfusion and then returned to its preligation level. A low level of ACH-PVIR lasted during the entire 120-min reperfusion. Similar changes in the ratio of the PVIR before and after treatment with NG or ACH were observed. In the 60-min group, capillary endothelial edema and widening of intercellular linking gaps were observed. CONCLUSIONS: We observed microvascular endothelial damage and endothelium-dependent dilatation impairments in stunned myocardium, and their severity and recovery rate are affected by the duration of ischemia.

Role of p-ERK1/2 in Benign Prostatic Hyperplasia during Hyperinsulinemia.

PURPOSE: Using a rat model of hyperinsulinemia, the present study investigated the role of p-ERK1/2 in benign prostatic hyperplasia (BPH). MATERIALS AND METHODS: Forty male Sprague-Dawley rats were randomly selected and assigned to four groups: high fat diet (HFD)+BPH (n=10), HFD (n=10), BPH (n=10), and control (n=10) groups. Hyperinsulinemia was induced by HFD feeding, while BPH was induced using testosterone propionate. Plasma glucose, plasma insulin and bodyweight were examined weekly. Immunohistochemistry (IHC) and western blot analysis were used to analyze the expression of ERK1/2 and p-ERK1/2 in rat prostates. RESULTS: Plasma glucose and plasma insulin levels were significantly greater in the HFD+BPH and HFD groups, when compared to the other two groups (P<0.05). Prostate weights were significantly greater in the HFD+BPH, HFD and BPH groups, than in the control group (P<0.05). IHC and western blot analysis revealed that p-ERK1/2 expression was greater in the HFD+BPH group than in the other three groups (P<0.05). CONCLUSION: Androgens plus a hyperinsulinemic condition induced by HFD can result in prostatic cell hyperplasia, and this mechanism may be correlated to the upregulation of p-ERK1/2. Further investigations of this possibility are required.

Boosting the biocatalytic precipitation with enzyme-loaded liposomes: Toward a general platform for amplified photoelectrochemical immunoassay.

Liposome-assisted photoelectrochemical (PEC) bioanalysis represents one of the latest frontiers in the arena of PEC bioanalysis. This work reports a general enzyme-amplified liposomal PEC bioanalysis protocol via the use of enzyme-loaded liposomes to boost the biocatalytic precipitation (BCP) effect. In the representative system, the horseradish peroxidase (HRP)-loaded liposome (HRPLL) and the Au nanoclusters (NCs)/Au nanoparticles (NPs)/TiO2 nanotubes (NTs) framework (AATF) were used as liposomal label and photoelectrode, respectively. In the detection, the sandwich immunocomplex reaction was accomplished in a 96-well plate to confine the HRPLL label, which was then lysed to release the HRP molecules to initiate the BCP process. Due to the amplified formation of HRP-induced BCP on the AATF scaffold, the photo-current response correlated closely with the immunorecognition process and the analyte could be detected very sensitively. This work features the first integration of enzyme-loaded liposomes and the BCP for sensitive PEC bioanalysis, which to our knowledge has not been reported. With the use of various other enzymes, this work could serve as a general basis for the PEC bioanalysis of numerous other target of interest.

[Evaluation of left ventricular diastolic function using velocity vector imaging and quantitative tissue velocity imaging].

OBJECTIVE: To validate the efficacy of velocity vector imaging (VVI) and quantitative tissue velocity imaging (QTVI) for evaluating left ventricular diastolic function. METHODS: Fifty-one patients underwent left heart catheterization were included in this study. Mean of peak early diastolic velocity (Em), EF and the ratio of early (E) to late (A) mitral valve flow velocity (E/A) were measured by echocardiography and the ratio of E to Em (E/Em) was calculated. Left ventricular end diastolic pressure (LVEDP) was measured during catheterization examination. RESULTS: E/Em derived from VVI or QTVI was significantly correlated with LVEDP (r = 0.808, P < 0.01 and r = 0.692, P < 0.01, respectively) and the correlation coefficient between VVI and LVEDP was significantly higher than that between QTVI and LVEDP (Z = 2.246, P = 0.025). Em derived from VVI and QTVI also negatively correlated with LVEDP (r = -0.740, P < 0.01 and r = -0.567, P < 0.01) and the correlation coefficient between VVI and LVEDP was significantly higher than that between QTVI and LVEDP (Z = 2.595, P = 0.009). However, there was no correlation between E/A and LVEDP (r = 0.117, P = 0.415). CONCLUSION: E/Em and Em derived from VVI and QTVI are valuable parameters for evaluating LV diastolic function.

Unique Redox Reaction between CuO Photocathode and Cysteine: Insight into the Mechanism for Cathodic Photoelectrochemical Bioanalysis.

Previously reported copper oxide-based cathodic photoelectrochemical bioanalysis of cysteine had attributed the decrease of the photocurrents to the binding of cysteine onto the CuO surface. However, our latest investigation found that the previous explanation was not correct. This Letter presents the in-depth study of such phenomena and a new insight into the underlying mechanism. Specifically, the unique redox reaction between the CuO photocathode and cysteine produced [Cys-Cu(I)] and cystine, and the insoluble complex blocked the partial contact between the photoelectrode and the dissolved O2-containing electrolyte and reduced the effective working area of the photocathode, leading to the decrease of photocurrent.

In situ chemical redox and functionalization of graphene oxide: toward new cathodic photoelectrochemical bioanalysis.

This report outlines the first exploration of graphene oxide (GO) itself as a light harvesting material with an innovative in situ chemical redox and functionalization (CRF) strategy for versatile and high-throughput cathodic photoelectrochemical (PEC) bioanalysis.