Covalent Coupling Assisted Hydrophilic Perovskite Spheres for Ratiometric Fluorescent Visual Multichannel Immunoassay.

Quantitative fluorescence immunoassay is essential for the construction of biosensing mechanisms and the quantification of trace markers. But the interference problems caused by low fluorescence efficiency and broad fluorescence spectrum of fluorescent probes have hindered the continued development of ratiometric fluorescence sensing in biosensing. Perovskite materials, with ultra-high color purity (FWHM < 30 nm) and photoluminescence quantum yield (PLQY) (close to 100%), are expected to be next-generation fluorescent probes. However, poor water stability and biocompatibility are still non-negligible in biosensor applications. In this work, hyperstatic perovskite fluorescent microspheres prepared by swelling-shrinking method can be used as ratiometric fluorescence signals and biological immunoassay platforms. Meanwhile, inspired by p-aminophenol (AP) controlled synthesis and the catalytic reaction of 4-aminophenol phosphate (APP) triggered by alkaline phosphatase (ALP), a strategy to prepare fluorescent nanoparticles as fluorescence signals for ALP detection is proposed. Most importantly, it is proposed for the first time to combine this enzymatic fluorescence with perovskite materials using covalent linkage to create a novel cascade immunoassay and use it for quantitative and visualization determination of hepatitis B surface antigen (HBsAg) for application verification. These results indicate the biosensing potential of perovskite materials and provide a pathway for high sensitivity enzyme detection and enzyme triggered immune detection.

Ginsenosides and Polysaccharides from Ginseng Co-Fermented with Multi-Enzyme-Coupling Probiotics Improve In Vivo Immunomodulatory Effects.

The active components of ginseng, such as ginsenosides and polysaccharides, have high therapeutic value in treating cancer, decreasing obesity, and enhancing immunity. However, simple primary ginseng treatment cannot maximize this medicinal potential. Therefore, in this study, Panax ginseng was co-fermented with multi-enzyme-coupling probiotics to obtain a fermentation broth with higher levels of ginsenosides, polysaccharides, and probiotics. When compared to other treatment methods for cyclophosphamide-induced immunosuppression in mice, the results reveal that the P. ginseng fermentation broth treated with multi-enzyme-coupling probiotics could significantly improve the immune function of immunosuppressive mice and restore intestinal flora stability. Overall, this processing method will provide a novel strategy for promoting the application of ginseng and the relief of immunosuppression.

Bi-allelic MEI1 variants cause meiosis arrest and non-obstructive azoospermia.

Non-obstructive azoospermia (NOA) is characterized by the failure of sperm production due to testicular disorders and represents the most severe form of male infertility. Growing evidences have indicated that gene defects could be the potential cause of NOA via genome-wide sequencing approaches. Here, bi-allelic deleterious variants in meiosis inhibitor protein 1 (MEI1) were identified by whole-exome sequencing in four Chinese patients with NOA. Testicular pathologic analysis and immunohistochemical staining revealed that spermatogenesis is arrested at spermatocyte stage, with defective programmed DNA double-strand breaks (DSBs) homoeostasis and meiotic chromosome synapsis in patients carrying the variants. In addition, our results showed that one missense variant (c.G186C) reduced the expression of MEI1 and one frameshift variant (c.251delT) led to truncated proteins of MEI1 in in vitro. Furthermore, the missense variant (c.T1585A) was assumed to affect the interaction between MEI1 and its partners via bioinformatic analysis. Collectively, our findings provide direct genetic and functional evidences that bi-allelic variants in MEI1 could cause defective DSBs homoeostasis and meiotic chromosome synapsis, which subsequently lead to meiosis arrest and male infertility. Thus, our study deepens our knowledge of the role of MEI1 in male fertility and provides a novel insight to understand the genetic aetiology of NOA.

Novel dual-reference approach facilitates the activation mapping and catheter ablation of premature atrial complexes with non-pulmonary vein and non-superior vena cava origins.

AIMS: Activation mapping of premature atrial complexes (PACs) proves challenging due to interference by mechanical bumping and non-targeted ectopies. This study aims to compare the mapping efficacy, instant success, and long-term recurrence of catheter ablation for PACs with non-pulmonary vein (PV) and non-superior vena cava (SVC) origins between the novel dual-reference approach (DRA) and the routine single-reference approach (SRA) of mapping. METHODS AND RESULTS: Patients with symptomatic, drug-refractory PACs, or frequent residual PACs after atrial tachyarrhythmia ablation were enrolled. During activation mapping, the coronary sinus (CS) catheter was used as the only timing reference in the SRA group. In the DRA group, another catheter, which was spatially separated from the CS catheter, was used as the second reference. The timing difference between the two references was used to discriminate the targeted PACs from the uninterested rhythms. Procedural parameters and long-term recurrence were compared. A total of 188 patients (109 in SRA and 79 in DRA) were enrolled. The baseline characteristics were similar. Compared with the SRA group, the DRA group had less repeated mapping (1.2 ± 0.4 vs. 1.4 ± 0.5, P = 0.004), shorter mapping (15 ± 6 vs. 23 ± 7 min, P < 0.001) and procedural time (119 ± 28 vs. 132 ± 22 min, P = 0.001), similar procedural complication rates (3.6 vs. 3.8%, P > 0.999), higher instant success (96.2 vs. 87.2%, P = 0.039), and lower recurrence rate (15.2 vs. 29.3%, hazard ratio 1.943, P = 0.033) during a 24-month follow-up. CONCLUSION: As a novel strategy, the DRA shortens the procedural time and improves both instant and long-term success of PAC ablation, serving as a promising approach in mapping PACs with non-PV and non-SVC origins.

The feasibility of proteomics sequencing based immune-related prognostic signature for predicting clinical outcomes of bladder cancer patients.

BACKGROUND: Bladder cancer (BCa) shows its potential immunogenity in current immune-checkpoint inhibitor related immunotherapies. However, its therapeutic effects are improvable and could be affected by tumor immune microenvironment. Hence it is interesting to find some more prognostic indicators for BCa patients concerning immunotherapies. METHODS: In the present study, we retrospect 129 muscle-invasive BCa (MIBC) patients with radical cystectomy in Shanghai General Hospital during 2007 to 2018. Based on the results of proteomics sequencing from 9 pairs of MIBC tissue from Shanghai General Hospital, we focused on 13 immune-related differential expression proteins and their related genes. An immune-related prognostic signature (IRPS) was constructed according to Cancer Genome Atlas (TCGA) dataset. The IRPS was verified in ArrayExpress (E-MTAB-4321) cohort and Shanghai General Hospital (General) cohort, separately. A total of 1010 BCa patients were involved in the study, including 405 BCa patients in TCGA cohort, 476 BCa patients in E-MTAB-4321 cohort and 129 MIBC patients in General cohort. RESULT: It can be indicated that high IRPS score was related to poor 5-year overall survival and disease-free survival. The IRPS score was also evaluated its immune infiltration. We found that the IRPS score was adversely associated with GZMB, IFN-γ, PD-1, PD-L1. Additionally, higher IRPS score was significantly associated with more M2 macrophage and resting mast cell infiltration. CONCLUSION: The study revealed a novel BCa prognostic signature based on IRPS score, which may be useful for BCa immunotherapies.

pH-induced highly sensitive fluorescence detection of urea and urease based on carbon dots-based nanohybrids.

Carbon quantum dots (CDs) have become one of the most popular fluorescent materials due to their intriguing performance, which are favored by many fields. However, it is difficult to synthesize CDs with high quantum yield by the simple synthesis methods. In this paper, we fabricated CDs- silicon (SiO2) spheres composites via a versatile hydrothermal route. The prepared BCD-SiO2 composites exhibited an approximately 10-fold increase in the fluorescence intensity over that of BCDs. At the same time, the purification path was simplified by the facile separation of SiO2 spheres. The prepared BCD-SiO2 composites were used to fabricate a special sensing platform for the ultrasensitive detection of urea and urease, with detection limits of 1.67 µM and 0.002 mg/mL, respectively. Furthermore, this strategy was successfully applied to the detection of real samples. This result shows that as-prepared BCDs-SiO2 composites are promising for broad application to biological analysis.

Renal Denervation by Noninvasive Stereotactic Radiotherapy Induces Persistent Reduction of Sympathetic Activity in a Hypertensive Swine Model.

Background We have previously reported the feasibility of noninvasive stereotactic body radiotherapy (SBRT) as a novel approach for renal denervation. Methods and Results Herein, from a translational point of view, we assessed the antihypertensive effect and chronological evolution of SBRT-induced renal nerve injury within 6 months in a hypertensive swine model. Hypertension was induced in swine by subcutaneous implantation of deoxycorticosterone acetate pellets in combination with a high-salt diet. A single dose of 25 Gy with SBRT was delivered for renal denervation in 9 swine within 3.4±1.0 minutes. Blood pressure levels at baseline and 1 and 6 months post-SBRT were comparable to control (n=5), whereas renal norepinephrine was significantly lower at 6 months (P<0.05). Abdominal computed tomography, performed before euthanasia and renal function assessment, remained normal. Standard semiquantitative histological assessment showed that compared with control (1.4±0.4), renal nerve injury was greater at 1 month post-SBRT (2.3±0.3) and peaked at 6 months post-SBRT (3.2±0.8) (P<0.05), along with a higher proportion of active caspase-3-positive nerves (P<0.05). Moreover, SBRT resulted in continuous dysfunction of renal sympathetic nerves and low level of nerve regeneration in 6 months by immunohistochemistry analysis. Conclusions SBRT delivering 25 Gy for renal denervation was safe and related to sustained reduction of sympathetic activity by aggravating nerve damage and inhibiting nerve regeneration up to 6 months; however, its translation to clinical trial should be cautious because of the negative blood pressure response in the deoxycorticosterone acetate-salt hypertensive swine model.

PD-1-Positive Tumor-Associated Macrophages Define Poor Clinical Outcomes in Patients With Muscle Invasive Bladder Cancer Through Potential CD68/PD-1 Complex Interactions.

Tumor-associated macrophages (TAMs) regulate tumor immunity. Previous studies have shown that the programmed cell death protein 1 (PD-1)-positive TAMs have an M2 macrophage phenotype. CD68 is a biomarker of TAMs and is considered to be a poor prognostic marker of several malignancies. Our results show that PD-1-positive TAMs can be a negative survival indicator in patients with muscle-invasive bladder cancer (MIBC), and that the mechanistic effects could result due to a combination of PD-1 and CD68 activity. We analyzed 22 immune cell types using data from 402 patients with MIBC from the TCGA database, and found that a high immune score and M2 TAMs were strongly associated with poor clinical outcomes in patients with MIBC. Further, we analyzed resected samples from 120 patients with MIBC and found that individuals with PD-1-positive TAMs showed a reduction in 5-year overall survival and disease-free survival. Additionally, PD-1-positive TAMs showed a significant association with higher programmed death-ligand 1 (PD-L1) expression, the Ki67 index, the pT stage and fewer CD8-positive T cells. Through the co-immunoprecipitation (co-IP) assay of THP-1 derived macrophages, we found that CD68 can bind to PD-1. The binding of CD68 and PD-1 can induce M2 polarization of THP-1 derived macrophages and promote cancer growth. The anti-CD68 treatment combined with peripheral blood mononuclear cells (PBMC) showed obvious synergy effects on inhibiting the proliferation of T24 cells. Together, these results indicate for the first time that CD68/PD-1 may be a novel target for the prognosis of patients with MIBC.

Ubiquitin specific peptidase 5 promotes ovarian cancer cell proliferation through deubiquitinating HDAC2.

Globally, epithelial ovarian cancer (EOC) is the most common gynecological malignancy with poor prognosis. The expression and oncogenic roles of ubiquitin specific peptidase 5 (USP5) have been reported in several cancers except EOC. In the current study, USP5 amplification was highly prevalent in patients with EOC and associated with higher mRNA expression of USP5. USP5 amplification and overexpression was positively correlated with poor prognosis of patients of ovarian serous carcinomas. Disruption of USP5 profoundly repressed cell proliferation by inducing cell cycle G0/G1 phase arrest in ovarian cancer cells. Additionally, USP5 knockdown inhibited xenograft growth in nude mice. Knockdown of USP5 decreased histone deacetylase 2 (HDAC2) expression and increased p27 (an important cell cycle inhibitor) expression in vitro and in vivo. The promoting effects of USP5 overexpression on cell proliferation and cell cycle transition, as well as the inhibitory effects of USP5 overexpression on p27 expression were mediated by HDAC2. Moreover, USP5 interacted with HDAC2, and disruption of USP5 enhanced the ubiquitination of HDAC2. HDAC2 protein was positively correlated USP5 protein, and negatively correlated with p27 protein in ovarian serous carcinomas tissues. Collectively, our data suggest the oncogenic function of USP5 and the potential regulatory mechanisms in ovarian carcinogenesis.

Catheter ablation of ventricular arrhythmias originating from the para-Hisian region with reversed C-curve technique.

AIMS: Ventricular arrhythmias (VAs) originating from the para-Hisian region represent a challenging location. The long-term success rate of catheter ablation above the septal leaflet of the tricuspid valve is not ideal. This study aimed to investigate the safety and efficacy of catheter ablation for para-Hisian VAs via a direct approach under the septal valve with reversed C-curve technique. METHODS AND RESULTS: Twenty-five consecutive patients with para-Hisian VAs were included. Systematic mapping was performed in the right ventricle septum, including both the regions above and under the septal valve. Radiofrequency (RF) ablation was preferentially performed under the valve with reversed C-curve technique in all patients. If the ablation failed under the valve, it was then performed above the valve and even in aortic sinus cusps. The earliest ventricular activation preceding surface QRS (V-QRS) under the valve was significantly larger than that above the valve (34.8 ± 5.3 vs 27.8 ± 5.7 ms, P < .01). RF ablation under the valve with reversed C-curve technique achieved acute success in 22 of 25 (88%) patients. Junctional rhythm developed during ablation in 3 of 25 (12%) patients and no atrioventricular block occurred. In the remaining three patients, RF application above the valve failed to eliminate the VAs and one of them achieved successful ablation in the right coronary cusp. During a mean follow-up of 17.8 ± 9.4 months, no patients presented with VAs recurrence and no postprocedure complications occurred. CONCLUSIONS: Catheter ablation under the valve with reversed C-curve technique shows to be effective and safe for para-Hisian VAs.

Low serum gastrin associated with ER+ breast cancer development via inactivation of CCKBR/ERK/P65 signaling.

BACKGROUND: Gastrin is an important gastrointestinal hormone produced primarily by G-cells in the antrum of the stomach. It normally regulates gastric acid secretion and is implicated in a number of human disease states, but how its function affects breast cancer (BC) development is not documented. The current study investigated the suppressive effects of gastrin on BC and its underlying mechanisms. METHODS: Serum levels of gastrin were measured by enzyme-linked immunosorbent assay (ELISA) and correlation between gastrin level and development of BC was analyzed by chi-square test. Inhibitory effects of gastrin on BC were investigated by CCK-8 assay and nude mice models. Expressions of CCKBR/ERK/P65 in BC patients were determined through immunohistochemistry (IHC) and Western blot. Survival analysis was performed using the log-rank test. RESULTS: The results indicated that the serum level of gastrin in BC patients was lower compared with normal control. Cellular and molecular experiments indicated that reduction of gastrin is associated with inactivation of cholecystokinin B receptor (CCKBR)/ERK/P65 signaling in BC cells which is corresponding to molecular type of estrogen receptor (ER) positive BC. Furthermore, we found that low expression of gastrin/CCKBR/ERK /P65 was correlated to worse prognosis in BC patients. Gastrin or ERK/P65 activators inhibited ER+ BC through CCKBR-mediated activation of ERK/P65. Moreover, combination treatment with gastrin and tamoxifen more efficiently inhibited ER+ BC than tamoxifen alone. CONCLUSIONS: We concluded that low serum gastrin is related to increased risk of ER+ BC development. The results also established that CCKBR/ERK/P65 signaling function is generally tumor suppressive in ER+ BC, indicating therapies should focus on restoring, not inhibiting, CCKBR/ERK/P65 pathway activity.

Prognostic and predictive values of immune infiltrate in patients with head and neck squamous cell carcinoma.

This study sought to determine whether the in situ tumor-infiltrating immune lymphocytes, as a novel companion to the Immunoscore analysis, could be a promising, valuable prognostic and predictive marker in patients with head and neck squamous cell carcinoma (HNSCC). Total (CD3+) and cytotoxic (CD8+) T lymphocytes were assessed using immunohistochemistry in tumor nests and stroma obtained from patient surgical specimens. The "Immunoscore" methodology has been defined to quantify the amount of in situ immune infiltrate (from I0 to I4). Survival curves were measured using the Kaplan-Meier method, and differences in survival and response to therapy between the groups were estimated using the log-rank test. The prognostic value of the Immunoscore was determined using Cox multivariate analysis. The density and location of CD3+ and CD8+ lymphocytes and the associated Immunoscore correlated significantly with differences in disease-free survival (DFS) and overall survival (OS) (all P < .005). Compared with tumor-node-metastasis (TNM) staging, the Immunoscore was found to have an advantage in predicting survival (P = .000). In addition, a high Immunoscore was associated with the tumors of advanced-stage patients who underwent different treatment regimens. The Immunoscore could be a useful prognostic marker. The measurement of CD3+ and CD8+ cell infiltration may be beneficial in HNSCC patients and may help determine which patients may benefit most from definitive chemoradiotherapy.

A novel P700 redox kinetics probe for rapid, non-intrusive and whole-tissue determination of photosystem II functionality, and the stoichiometry of the two photosystems in vivo.

We sought a rapid, non-intrusive, whole-tissue measure of the functional photosystem II (PS II) content in leaves. Summation of electrons, delivered by a single-turnover flash to P700(+) (oxidized PS I primary donor) in continuous background far-red light, gave a parameter S in absorbance units after taking into account an experimentally determined basal electron flux that affects P700 redox kinetics. S was linearly correlated with the functional PS II content measured by the O(2) yield per single-turnover repetitive flash in Arabidopsis thaliana expressing an antisense construct to the PsbO (manganese-stabilizing protein in PS II) proteins of PS II (PsbO mutants). The ratio of S to z(max) (total PS I content in absorbance units) was comparable to the PS II/PS I reaction-center ratio in wild-type A. thaliana and in control Spinacea oleracea. Both S and S/z(max) decreased in photoinhibited spinach leaf discs. The whole-tissue functional PS II content and the PS II/photosystem I (PS I) ratio can be non-intrusively monitored by S and S/z(max), respectively, using a quick P700 absorbance protocol compatible with modern P700 instruments.

Pathobiology of cancer chemotherapy-induced peripheral neuropathy (CIPN).

Chemotherapy induced peripheral neuropathy (CIPN) is a type of neuropathic pain that is a major dose-limiting side-effect of potentially curative cancer chemotherapy treatment regimens that develops in a "stocking and glove" distribution. When pain is severe, a change to less effective chemotherapy agents may be required, or patients may choose to discontinue treatment. Medications used to alleviate CIPN often lack efficacy and/or have unacceptable side-effects. Hence the unmet medical need for novel analgesics for relief of this painful condition has driven establishment of rodent models of CIPN. New insights on the pathobiology of CIPN gained using these models are discussed in this review. These include mitochondrial dysfunction and oxidative stress that are implicated as key mechanisms in the development of CIPN. Associated structural changes in peripheral nerves include neuronopathy, axonopathy and/or myelinopathy, especially intra-epidermal nerve fiber (IENF) degeneration. In patients with CIPN, loss of heat sensitivity is a hallmark symptom due to preferential damage to myelinated primary afferent sensory nerve fibers in the presence or absence of demyelination. The pathobiology of CIPN is complex as cancer chemotherapy treatment regimens frequently involve drug combinations. Adding to this complexity, there are also subtle differences in the pathobiological consequences of commonly used cancer chemotherapy drugs, viz platinum compounds, taxanes, vincristine, bortezomib, thalidomide and ixabepilone, on peripheral nerves.