Conjugation of sulpiride with a cell penetrating peptide to augment the antidepressant efficacy and reduce serum prolactin levels.

Depression ranks as the fourth most prevalent global disease, with suicide incidents occurring at a younger age. Sulpiride (SUL), an atypical antidepressant drug acting as a dopamine D2 receptor antagonist and possessing anti-inflammatory properties, exhibits limited ability to penetrate the blood brain barrier (BBB). This weak penetration hampers its inhibitory effect on prolactin release in the pituitary gland, consequently leading to hyperprolactinemia. In order to enhance the central nervous system efficacy of sulpiride and reduce serum prolactin levels, we covalently linked sulpiride to VPALR derived from the nuclear DNA repair protein ku70. In vivo study on depressive mice using intraperitoneal injection of VPALR-SUL demonstrated a significant increase in struggle time and total distance compared to those treated with only sulpiride while also reducing serum prolactin concentration. The pharmacokinetic study results showed that VPALR-SUL prolonged half-life and increased bioavailability. In conclusion, VPALR-SUL exhibited potential for enhancing sulpiride transport across the BBB, augmenting its antidepressant effects, and reducing serum prolactin levels. This study laid a foundation for improving sulpiride delivery and developing novel antidepressants.

A sensitive and specific LC-MS/MS method for determination of a novel antihypertensive peptide FR-6 in rat plasma and pharmacokinetic study.

The investigation of peptide drugs has become essential in the development of innovative medications for hypertension. In this study, a sensitive high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method was developed to determine the plasma concentration and stability of the antihypertensive peptide FR-6 in rats. An isotopically labeled peptide (with an unchanged sequence) was utilized as an internal standard (IS) for validation purposes. Subsequently, this assay was employed to examine the pharmacokinetics of different administration methods (tail vein and gavage) in Sprague Dawley (SD) rats. Extracted plasma samples underwent sample preparation through methanol protein precipitation, followed by elution of FR-6 on Wondasil C18 Superb column (4.6 × 150 mm, 5 µm), using a mobile phase consisting of formic acid (0.1%) in water (A) and formic acid (0.125%)-ammonium formate (2 mM) in methanol (B). Ion pairs corresponding to FR-6 and IS were monitored via multiple reaction monitoring (MRM) under positive ion mode: m/z 400.7 â†’ 285.1 for FR-6 and m/z 406.1 â†’ 295.1 for IS detection respectively. The method exhibited excellent linearity with respect to FR-6 concentrations. In addition, the inter-day and intra-day precision were 0.61-6.85% and 1.76-11.75%; the inter-day and intra-day accuracy were -7.28-0.13% and -7.20-2.28%, respectively. In conclusion, the matrix effect, extraction recovery, and stability data were validated according to FDA recommended acceptance criteria for bioanalytical methods. This validated method serves as a reliable tool for determining the concentration of antihypertensive peptide FR-6, and has been successfully applied in pharmacokinetic studies involving rats.

Flux optimization using multiple promoters in Halomonas bluephagenesis as a model chassis of the next generation industrial biotechnology.

Predictability and robustness are challenges for bioproduction because of the unstable intracellular synthetic activities. With the deeper understanding of the gene expression process, fine-tuning has become a meaningful tool for biosynthesis optimization. This study characterized several gene expression elements and constructed a multiple inducible system that responds to ten different small chemical inducers in halophile bacterium Halomonas bluephagenesis. Genome insertion of regulators was conducted for the purpose of gene cluster stabilization and regulatory plasmid simplification. Additionally, dynamic ranges of the multiple inducible systems were tuned by promoter sequence mutations to achieve diverse scopes for high-resolution gene expression control. The multiple inducible system was successfully employed to precisely control chromoprotein expression, lycopene and poly-3-hydroxybutyrate (PHB) biosynthesis, resulting in colorful bacterial pictures, optimized cell growth, lycopene and PHB accumulation. This study demonstrates a desirable approach for fine-tuning of rational and efficient gene expressions, displaying the significance for metabolic pathway optimization.

The Prognostic Significance of the TEAD4 in Hepatocellular Carcinoma.

Background: Abnormal expression of genes causes tumorigenesis, tumor progression, and poor prognosis in hepatocellular carcinoma (HCC). Therefore, the aims of this study were to explore the transcription enhancer domain factor 4 (TEAD4) in patients with liver cancer and its relationship with prognosis. Methods: HTSeq-FPKM data and corresponding clinical data of HCC patients were obtained from The Cancer Genome Atlas (TCGA). Difference in TEAD4 expression between normal and tumor and the correlation with clinical characteristics were analyzed by the chi-squared test based on UALCAN. HepG2 cell lines were used to study the effect of TEAD4 on HCC cell lines. The expression and clinical significance of TEAD4 in HCC were detected in clinical cases. Results: The transcription and post-transcription levels of TEAD4 were higher in HCC tumors than normal illustrated different expressed transcription of TEAD4 in gender, nodal metastasis status, tumor grades, and individual cancer stages. The high TEAD4 expression was significantly associated with tumor grades. The high expression of TEAD4 was significantly correlated to shorter 2-5 years overall survival. Inhibition of TEAD4 expression in HepG2 cells resulted in significantly decreased cell proliferation and invasion. Conclusion: TEAD4 was identified as an independent prognostic factor, and inhibition of TEAD4 expression in HepG2 cells resulted in significantly decreased cell proliferation and invasion.

Phase separation of α-crystallin-GFP protein and its implication in cataract disease.

Cataract, the leading cause of blindness worldwide, is caused by crystallin protein aggregation within the protected lens environment. Phase separation has been implicated as an important mechanism of protein aggregation diseases, such as neurodegeneration. Similarly, cataract has been proposed to be a protein condensation disease in the last century. However, whether crystallin proteins aggregate via a phase separation mechanism and which crystallin protein initiates the aggregation remain unclear. Here, we showed that all types of crystallin-GFP proteins remain soluble under physiological conditions, including protein concentrations, ion strength, and crowding environments. However, in age or disease-induced aberrant conditions, α-crystallin-GFP, including αA- and αB-crystallin-GFP, but not other crystallin-GFP proteins, undergo phase separation in vivo and in vitro. We found that aging-related changes, including higher crystallin concentrations, increased Na+, and decreased K+ concentrations, induced the aggregation of α-crystallin-GFP. Furthermore, H2O2, glucose, and sorbitol, the well-known risk factors for cataract, significantly enhanced the aggregation of αB-crystallin-GFP. Taken together, our results revealed that α-crystallin-GFP forms aggregates via a phase transition process, which may play roles in cataract disease. Opposite to the previously reported function of enhancing the solubility of other crystallin, α-crystallin may be the major aggregated crystallin in the lens of cataract patients.

A novel and sensitive ratiometric fluorescent quantum dot-based biosensor for alkaline phosphatase detection in biological samples via the inner-filter effect.

Alkaline phosphatase (ALP) is an important biomarker whose abnormal level in activity is associated with hepatobiliary, skeletal, and renal diseases as well as cancer. Herein, we synthesized ZnSe@ZnS quantum dots (ZnSe@ZnS QDs) and Mn-doped ZnS quantum dots (Mn:ZnS QDs) as fluorophores to establish the ratiometric fluorescent assay for ALP activity detection in biological samples. p-Nitrophenyl phosphate (PNPP) was used as a substrate for ALP, and the overlaps between absorption spectra of PNPP and excitation spectra of QDs resulted in sharp fluorescence quenching. Under the catalysis of ALP, PNPP was hydrolyzed into p-nitrophenol (PNP), which caused a red shift of absorption band of PNPP and fluorescence recovery of Mn:ZnS QDs (585 nm). However, the overlaps between absorption spectra of PNP and emission spectra of ZnSe@ZnS QDs led a further quenching of ZnSe@ZnS QDs (405 nm). Therefore, the ratiometric fluorescent signals (F 585/F 405) were associated with activity of ALP based on bidirectional responses of QDs to the concentration of PNPP. Under the optimum conditions, the method exhibited a good linear relationship from 4 to 96 U per L (R 2 = 0.9969) with the detection limit of 0.57 U per L. Moreover, the method was successfully applied for detecting the ALP activity in a complex biological matrix (human serum and HepG2 cells) with impressive specificity. In particular, the complicated chemical modifications of QDs and pretreatments of biological samples were not required in the whole detection procedures. Therefore, it not only provided a sensitive, specific and simple approach to clinical ALP activity detection, but it also provided support for early diagnosis of diseases.

MicroRNA-155-5p in serum derived-exosomes promotes ischaemia-reperfusion injury by reducing CypD ubiquitination by NEDD4.

AIMS: Recovery of blood flow is a therapeutic approach for myocardial infarction but paradoxically induces injury to the myocardium. Exosomes (exos) are pivotal mediators for intercellular communication that can be released by different cells and are involved in cardiovascular diseases. This study aimed to explore the possible effects and mechanisms of miR-155-5p loaded by serum-derived exos in myocardial infarction reperfusion injury (MIRI). METHODS AND RESULTS: Exos were isolated from mouse serum after induction of ischaemia reperfusion (I/R) and injected into I/R-treated mice to assess cardiac function, infarction size, and cardiomyocyte apoptosis. Primary cardiomyocytes were transfected with miR-155-5p inhibitor before treatment with oxygen-glucose deprivation and re-oxygenation (OGD/R) and exos derived from the serum of I/R-treated mice (I/R-Exos), in which Bcl-2, Bax, and cleaved-caspase-3 levels were detected. The interactions among miR-155-5p, NEDD4, and CypD were evaluated. miR-155-5p level was evidently increased in I/R-Exos than in exos from the serum of sham-operated mice (P < 0.05). In comparison with the I/R group, the I/R-Exos + I/R group had increased infarct size, elevated miR-155-5p expression, and boosted apoptotic rate in mouse myocardium (P < 0.05). In mice treated with I/R-Exos and I/R, miR-155-5p inhibition reduced cardiac infarct size and apoptosis (P < 0.05). NEDD4 was a target gene of miR-155-5p and promoted CypD ubiquitination. Cardiomyocyte apoptosis was markedly increased in the miR-155-5p inhibitor + shNEDD4 + OGD/R group versus the miR-155-5p inhibitor + OGD/R group (P < 0.05), but decreased in the miR-155-5p inhibitor + shNEDD4 + shCypD + OGD/R group than in the miR-155-5p inhibitor + shNEDD4 + OGD/R group (P < 0.05). CONCLUSIONS: miR-155-5p in I/R-Exos may facilitate MIRI by inhibiting CypD ubiquitination via targeting NEDD4.

The Relationship between Sleeping Position and Sleep Quality: A Flexible Sensor-Based Study.

The use of flexible wearable sensors to monitor the impact of sleeping position and turning frequency on sleep and to study sleep patterns can help bedridden patients heal and recover. The flexible wearable sleeping-position monitoring device was designed and developed using a flexible angle sensor and a six-axis motion sensor to measure the dynamic changes in body posture during sleep. Based on the changes in the output parameters of the flexible angle sensor and the six-axis motion sensor, we determined the change in the subject's lying position, verifying and analyzing the relationship between lying position preference, turning frequency, and sleep quality in healthy subjects. The sleeping-position monitoring device was worn by 13 subjects (7 males and 6 females) without sleep disorders before the sleep experiment. They performed more than 50 sleeping-position changes to ensure the accuracy of the monitoring device. Subjects slept in their beds for 8 h per night for 15 nights. During that time, they wore the sleeping-position monitoring device and a wristband sleep-monitoring bracelet on their left hand, and gathered the subjective sleep data using questionnaires. The results show that the most critical influencing factors are sleeping-position preference and frequency of turning. Data analysis reveals that subjects with a preference for right-sided lying and a lower frequency of turning had better sleep quality.

A Pipeline for Predicting the Treatment Response of Neoadjuvant Chemoradiotherapy for Locally Advanced Rectal Cancer Using Single MRI Modality: Combining Deep Segmentation Network and Radiomics Analysis Based on "Suspicious Region".

Patients with locally advanced rectal cancer (LARC) who achieve a pathologic complete response (pCR) after neoadjuvant chemoradiotherapy (nCRT) typically have a good prognosis. An early and accurate prediction of the treatment response, i.e., whether a patient achieves pCR, could significantly help doctors make tailored plans for LARC patients. This study proposes a pipeline of pCR prediction using a combination of deep learning and radiomics analysis. Taking into consideration missing pre-nCRT magnetic resonance imaging (MRI), as well as aiming to improve the efficiency for clinical application, the pipeline only included a post-nCRT T2-weighted (T2-w) MRI. Unlike other studies that attempted to carefully find the region of interest (ROI) using a pre-nCRT MRI as a reference, we placed the ROI on a "suspicious region", which is a continuous area that has a high possibility to contain a tumor or fibrosis as assessed by radiologists. A deep segmentation network, termed the two-stage rectum-aware U-Net (tsraU-Net), is designed to segment the ROI to substitute for a time-consuming manual delineation. This is followed by a radiomics analysis model based on the ROI to extract the hidden information and predict the pCR status. The data from a total of 275 patients were collected from two hospitals and partitioned into four datasets: Seg-T (N = 88) for training the tsraUNet, Rad-T (N = 107) for building the radiomics model, In-V (N = 46) for internal validation, and Ex-V (N = 34) for external validation. The proposed method achieved an area under the curve (AUC) of 0.829 (95% confidence interval [CI]: 0.821, 0.837) on In-V and 0.815 (95% CI, 0.801, 0.830) on Ex-V. The performance of the method was considerable and stable in two validation sets, indicating that the well-designed pipeline has the potential to be used in real clinical procedures.

Dihydropyrimidinase Like 2 Promotes Bladder Cancer Progression via Pyruvate Kinase M2-Induced Aerobic Glycolysis and Epithelial-Mesenchymal Transition.

BACKGROUND: Aerobic glycolysis and epidermal-mesenchymal transition (EMT) play key roles in the development of bladder cancer. This study aimed to investigate the function and the underlying mechanism of dihydropyrimidinase like 2 (DPYSL2) in bladder cancer progression. METHODS: The expression pattern of DPYSL2 in bladder cancer and the correlation of DPYSL2 expression with clinicopathological characteristics of bladder cancer patients were analyzed using the data from different databases and tissue microarray. Gain- and loss-of-function assays were performed to explore the role of DPYSL2 in bladder cancer progression in vitro and in mice. Proteomic analysis was performed to identify the interacting partner of DPYSL2 in bladder cancer cells. FINDINGS: The results showed that DPYSL2 expression was upregulated in bladder cancer tissue compared with adjacent normal bladder tissue and in more aggressive cancer stages compared with lower stages. DPYSL2 promoted malignant behavior of bladder cancer cells in vitro, as well as tumor growth and distant metastasis in mice. Mechanistically, DPYSL2 interacted with pyruvate kinase M2 (PKM2) and promoted the conversion of PKM2 tetramers to PKM2 dimers. Knockdown of PKM2 completely blocked DPYSL2-induced enhancement of the malignant behavior, glucose uptake, lactic acid production, and epithelial-mesenchymal transition in bladder cancer cells. INTERPRETATION: In conclusion, the results suggest that DPYSL2 promotes aerobic glycolysis and EMT in bladder cancer via PKM2, serving as a potential therapeutic target for bladder cancer treatment.

The activation of GPER inhibits cells proliferation, invasion and EMT of triple-negative breast cancer via CD151/miR-199a-3p bio-axis.

Background: Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype. G protein coupled receptor (GPER), the key player in the intercellular signaling communication, has been verified to participate in tumorigenesis. The present study aims to explore the effects of GPER on cell proliferation, invasion and EMT through CD151/miR-199a-3p bio-axis in TNBC cells. Methods: Total proteins were isolated from TNBC cell lines and GPER expression was determined using western blot assay. CCK-8 assay was used to detect cell viability after being treated with GPER activation. Western blotting and immunofluorescence were applied to measure the level of proteins associated with cell proliferation, angiogenesis and EMT, as well as the Hippo signal pathway. The level of miR-199a-3p and transfection efficiency were evaluated by reverse transcriptase quantitative PCR (RT-qPCR) after being transfected with miR-199a-3p mimics. Cell migration and invasion of TNBC cells were assessed by wound healing and transwell assays. Moreover, luciferase reporter assay was conducted to verify the relationship between CD151 and miR-199a-3p. Results: GPER activation treatment suppressed MDA-MB-231 cell viability, proliferation, migration, invasion, angiogenesis and EMT process. The expression of E-cadherin was increased, but N-cadherin, Vimentin, VEGFA, AngII and CD151 were decreased after GPER activation treatment. Conversely, inhibition of GPER indeed up-regulated CD151 expression. In addition, overexpression of miR-199a-3p supressed cell proliferation, migration, invasion and angiogenesis, as well as EMT process and the Hippo signal pathway. Conclusion: Collectively, the activation of GPER inhibits cells proliferation, invasion and EMT of triple-negative breast cancer via CD151/miR-199a-3p bio-axis. This study provides a novel intervention target for the treatment of breast cancer cells and a fresh idea for the clinical therapy of breast cancer.

Secreted TGF-beta-induced protein promotes aggressive progression in bladder cancer cells.

Background: Transforming growth factor-beta-induced (TGFBI) is an exocrine protein, which has been found to be able to promote the development of nasopharyngeal carcinoma, glioma, pancreatic cancer, and other tumors. However, there is currently no report concerning the relationship between TGFBI and invasive progression of bladder cancer (BCa). Methods: IHC staining, qRT-PCR and Western blot were used to analyze TGFBI and EMT markers levels. In vivo tumorigenesis was performed by xenograft tumor model. Results: In this study, we found that both mRNA and protein levels of TGFBI were significantly up-regulated in muscle invasive bladder cancer (MIBC) tissues compared with non-muscle-invasive bladder cancer (NMIBC) tissues. The high expression level of TGFBI was positively correlated with high histological grade and advanced clinical stage, and BCa patients with high TGFBI levels exhibited poor prognoses. We further confirmed that high expression level of TGFBI can promote proliferation, invasive progression, and epithelial-to-mesenchymal transition (EMT) of BCa cells in vitro, as well as promote tumor growth and EMT in vivo, while silencing of TGFBI inhibited these malignant phenotypes. TGFBI was involved in the up-regulation of EMT by inducing the expression level of Slug, Vimentin, Snail, MMP2, and MMP9 genes, while it down-regulated the expression level of E-cadherin. Moreover, Western blot analysis was carried out to demonstrate that BCa cell lines stably transfected with expression of TGFBI, a secreted protein. Furthermore, conditional medium containing TGFBI protein also resulted in enhanced EMT and malignant phenotype of BCa cells. Conclusion: Our results indicate that high expression level of TGFBI promotes EMT, proliferation, and invasive progression of BCa cells, and TGFBI is a potential therapeutic target and prognostic marker for BCa.

Overexpression of TPX2 is associated with progression and prognosis of prostate cancer.

Targeting protein for Xenopus kinesin-like protein 2 (TPX2) activates Aurora kinase A during mitosis and targets its activity to the mitotic spindle, serving an important role in mitosis. It has been associated with different types of cancer and is considered to promote tumor growth. The aim of the present study was to explore the role of TPX2 in diagnosing prostate cancer (PCa). It was identified that TPX2 expression in PCa tissues was increased compared with benign prostate tissues. Microarray analysis demonstrated that TPX2 was positively associated with the Gleason score, tumor-node-metastasis (TNM) stage, clinicopathological stage, metastasis, overall survival and biochemical relapse-free survival. In vitro studies revealed that the high expression of TPX2 in PCa cells improved proliferative, invasive and migratory abilities, and repressed apoptosis of the PCa cells, without affecting tolerance to docetaxel. The results suggested that TPX2 serves as a tumorigenesis-promoting gene in PCa, and a potential therapeutic target for patients with PCa.

Connexin43 interacts with Caveolin-3 in the heart.

Gap junctions (GJs), collections of multiple intercellular channels between neighboring cells, are specialized channels facilitating intercellular electrical and chemical communication. GJs are important for synchronizing coupling and coordinated contraction in the heart, and are crucial regulators of heart gene transcription, cardiac development, and protection of ischemic cardiomyocytes through second messenger communication. Identification of proteins that interact with Connexin43 (Cx43), the predominant protein in cardiac GJs, may contribute to the understanding of GJ functional regulation. Using a yeast two-hybrid system, we identified Caveolin-3 (Cav3) as a new Cx43-interacting protein. This interaction was confirmed by co-immunoprecipitation and co-localization experiments. CX43 interacts with Cav3, suggesting that Cav3 may participate in the functional regulation of GJs.