Rationale: In combination chemotherapy, the molar ratio of drugs is a critical parameter that determines the synergistic effects. However, most co-delivery vectors are incapable of maintaining the optimal molar ratio of drugs throughout the delivery process. Herein, a calixarene-modified albumin (CaMA), which can co-deliver multiple drugs with precise control of the drug ratio, is presented. Methods: CaMA was prepared by chemically conjugating multiple sulfonate azocalix[4]arenes (SAC4A) onto the surface of bovine serum albumin (BSA). The precise drug loading and synchronous drug release were measured using fluorescence spectroscopy. Mouse tumor cell 4T1 and 4T1-bearing mice were used to evaluate the combined effects of mitomycin C (MMC) and doxorubicin (DOX) in vitro and in vivo. Results: With multiple hypoxia-responsive calixarenes conjugated onto a single albumin molecule, CaMA achieved precise drug loading and synchronous release of multiple drugs into the tumor microenvironment. This unique drug loading and release mechanism ensures that CaMA maintains the drug ratio from the initial drug loading to the release site, providing a solid foundation for multi-drug combination therapy with the goal of achieving predictable therapeutic outcomes in vivo. The delivery of the model drug combination MMC and DOX at a prescreened ratio via CaMA achieved significantly enhanced tumor suppression and reduced systemic toxicity. Conclusions: This stoichiometric delivery feature makes CaMA a powerful tool for the development of combination chemotherapy and personalized medications for cancer treatment.

This study aimed to investigate the effects of diets supplemented with different levels of rosemary extract (RE) on the growth performance, meat quality, serum biochemistry, antioxidative capacity, and immunological capacity of Cherry Valley meat ducks. A total of 525 healthy Cherry Valley female meat ducks at 1 d of age were selected for this study. Ducks were randomly divided into 5 treatments with 7 replicates per treatment, and each replicate had 15 ducks. All replicates were randomly assigned to treatments. The study was designed as a dose response experiment. Treatment 1 (CON) was fed with the basal diet, and Treatment 2 to 5 (RE250, RE500, RE750, RE1000) were fed with the basal diet supplemented with 250, 500, 750, and 1,000 g/t RE, respectively. The whole experiment lasted 42 days with early stage (1-21 d) and late stage (22-42 d). Results showed that during 22 to 42 d, ducks that were fed over 500 g/t RE had significantly lower feed gain ratio than the ones in CON (P = 0.006). In addition, ducks in RE750 had significantly lower L* and a* in leg muscle compared with the ones in CON (P < 0.05). Besides, ducks that were fed between 250 and 750 g/t RE had significantly lower total protein level in serum compared with the ones in CON (P = 0.005). Ducks in RE250 and RE750 had significantly lower albumin, total cholesterol, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol levels in serum compared with the ones in CON and RE1000 (P < 0.05), and significant quadratic relationships were noticed between albumin, total cholesterol, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol and dietary RE level (P < 0.05). Moreover, ducks that were fed between 500 and 750 g/t RE had significantly higher levels of interleukin-2 in serum compared to the ones in CON and RE1000 (P = 0.003). Ducks in RE250 and RE750 had significantly higher levels of immunoglobulin G in serum compared to the ones in CON and RE1000 (P < 0.001). Ducks that were fed over 500 g/t RE had significantly higher levels of immunoglobulin A in serum compared to the ones in CON (P = 0.001). Finally, ducks that were fed between 500 and 750 g/t RE had significantly higher serum levels of glutathione peroxidase, superoxide dismutase, catalase, and total antioxidant capacity (P < 0.05) compared to the ones in CON. Ducks that were fed over 250 g/t RE had significantly lower serum level of malondialdehyde compared to the ones in CON (P = 0.020). Collectively, dietary supplementation of RE improved the growth performance and meat qualities of meat ducks during 22 to 42 d, which were possibly associated with the antioxidative and anti-inflammatory effects of RE. Based on the serum antioxidative and immunological parameters, we suggested that 500 to 750 g/t was the optimal supplementation rate for RE in diets for meat ducks aged 22 to 42 d.

Vibrio harveyi is regarded as serious pathogen for marine fishes. To evaluate the physiological responses of spotted sea bass (Lateolabrax maculatus) after V. harveyi infection, four biochemical biomarkers including alanine amino transferase (ALT), albumin (ALB), total protein (TP) and glucose (GLU) were measured in serum. Our results showed that V. harveyi infection significantly influenced the concentration of ALT, ALB and GLU. Additionally, five interleukin-17 (IL-17) and five IL-17 receptors (IL-17R) genes were identified in spotted sea bass and their gene structures were characterized. Furthermore, the expression patterns of IL-17 and IL-17R genes were determined by qPCR in liver, intestine, spleen and head kidney after V. harveyi infection. All IL-17 and IL-17R genes exhibited time- and tissue-dependent expressions. Several tested genes were dramatically induced by V. harveyi treatment, particularly IL-17A/F1 in liver and head kidney, IL-17A/F2 in head kidney, IL-17RC in spleen with more than 10-fold increases, which suggested their potential essential roles against bacterial infection.

In recent years, human serum albumin (HSA) has been characterized as an ideal drug carrier in the cancer arena. Caveolin-1 (Cav-1) has been established as the principal structural protein of caveolae and, thus, critical for caveolae-mediated endocytosis. Cav-1 has been shown to be overexpressed in cancers of the lung and pancreas, among others. We found that Cav-1 expression plays a critical role in both HSA uptake and response to albumin-based chemotherapies. As such, developing a novel albumin-based chemotherapy that is more selective for tumors with high Cav-1 expression or high levels of caveolar-endocytosis could have significant implications in biomarker-directed therapy. Herein, we present the development of a novel and effective HSA-SN-38 conjugate (SSH20). We find that SSH20 uptake decreases significantly by immunofluorescence assays and western blotting after silencing of Cav-1 expression through RNA interference. Decreased drug sensitivity occurs in Cav-1-depleted cells using cytotoxicity assays. Importantly, we find significantly reduced sensitivity to SSH20 in Cav-1-silenced tumors compared to Cav-1-expressing tumors in vivo. Notably, we show that SSH20 is significantly more potent than irinotecan in vitro and in vivo. Together, we have developed a novel HSA-conjugated chemotherapy that is potent, effective, safe, and demonstrates improved efficacy in high Cav-1-expressing tumors.

Current long term cryopreservation of cell stocks routinely requires the use of liquid nitrogen (LN2), because commonly used cryopreservation media containing cell membrane permeating cryoprotectants are thermally unstable when frozen at higher storage temperatures, e.g. -80 °C. This instability leads to ice recrystallization, causing progressive loss of cell viability over time under the storage conditions provided by most laboratory deep freezers. The dependency on LN2 for cell storage significantly increases operational expense and raises issues related to impaired working efficiency and safety. Here we demonstrate that addition of Ficoll 70 to cryoprotectant solutions significantly improves system thermal stability at the working temperature (~-80 °C) of laboratory deep freezers. Moreover, a medium comprised of Ficoll 70 and dimethyl sulfoxide (DMSO) in presence or absence of fetal bovine serum (FBS) can provide reliable cryopreservation of various kinds of human and porcine pluripotent stem cells at -80 °C for periods that extend up to at least one year, with the post-thaw viability, plating efficiency, and full retention of pluripotent phenotype comparable to that achieved with LN2 storage. These results illustrate the practicability of a promising long-term cryopreservation method that completely eliminates the need for LN2.

Traditional Chinese medicines are gaining more attention as promising adjuvant agents for conventional chemotherapy. Recent studies have shown that lobetyolin (LBT) is one of the main bioactive compounds of traditional Chinese medicines and it exhibits anticancer activity in several types of cancer. Therefore, this study aimed to investigate the mechanism by which LBT inhibits lung cancer. A549 human lung cancer cells were treated with LBT. In addition, A549 cells were injected into Balc/b nude mice to establish model of lung cancer. The mice were treated with cisplatin (DDP) or LBT alone or in combination, and tumor growth was monitored. Protein levels of E-cadherin, vimentin and matrix metalloproteinase 9 (MMP9) were detected. We found that the combination of LBT and DDP showed stronger effect to inhibit the proliferation of A549 cells compared to LBT or DDP treatment alone. Wound healing assay showed that the ratio of wound healing was significantly lower in LBT group and DDP group and was the lowest in LBT+DDP group. Transwell invasion assay showed that the invasion ability of A549 cells was the weakest in LBT+DDP group. Protein levels of E-cadherin were the highest while those of vimentin and MMP9 were the lowest in A549 cells treated with LBT+DDP. Nude mouse xenograft tumor model showed that the combination of LBT with DDP had the highest efficacy to inhibit the growth of lung cancer, and tumor tissues of mice treated with LBT+DDP had the lowest expression of vimentin and MMP9 and the highest expression of E-cadherin. In conclusion, LBT significantly enhances the efficacy of chemotherapy on lung cancer, and the mechanism may be related to the inhibition of epithelial-mesenchymal transition.

In spite of the tumor microenvironments responsive cancer therapy based on Fenton reaction (i.e., chemodynamic therapy, CDT) has been attracted more attentions in recent years, the limited Fenton reaction efficiency is the important obstacle to further application in clinic. Herein, we synthesized novel FeO/MoS2 nanocomposites modified by bovine serum albumin (FeO/MoS2-BSA) with boosted Fenton reaction efficiency by the synergistic effect of co-catalyze and photothermal effect of MoS2 nanosheets triggered by the second near-infrared (NIR II) light. In the tumor microenvironments, the MoS2 nanosheets not only can accelerate the conversion of Fe3+ ions to Fe2+ ions by Mo4+ ions on their surface to improve Fenton reaction efficiency, but also endow FeO/MoS2-BSA with good photothermal performances for photothermal-enhanced CDT and photothermal therapy (PTT). Consequently, benefiting from the synergetic-enhanced CDT/PTT, the tumors are eradicated completely in vivo. This work provides innovative synergistic strategy for constructing nanocomposites for highly efficient CDT.

Perivascular adipose tissue (PVAT), an active endocrine organ, exerts direct effect on vascular tone through paracrine. Activation of PVAT metabolism plays an inhibitory role in atherosclerosis via secreting relaxing factors. The present studies were designed to investigate the role of PVAT metabolism in regulation of hypertension.

Osteosarcoma is the most common malignancy of the bones, and although advances in chemotherapy and surgery had been achieved in recent years, the long-term survival rate has reached a plateau. The main reason for this is the aggressive malignant potential and poor response of the disease to chemotherapy. However, several studies have found that tumor resistance is associated with cancer stem cells (CSCs). To address this issue, in the present study, osteosarcoma cells were treated with specially designated concentrations of cisplatin (CDDP) in a mouse model. Hematoxylin and eosin staining analyses were performed to assess tissue structure, in vivo passaging and CDDP treatment. Drug resistance genes and well-established stemness genes were detected by quantitative polymerase chain reaction. A serum-starved sphere formation assay was adopted to evaluate the ability to generate spherical clones and flow cytometry as used to test the expression of the cluster of differentiation 117 and Stro-1 surface markers, known as markers of CSCs. It was found that CDDP could induce an effect of resistance in the osteosarcoma cells, which possessed cancer stem CSC properties, as shown by the elevated expression of CSC marker genes and the higher expression of the cluster of differentiation 117 and Stro-1 surface markers. Moreover, the cells that dissociated from the tumor tissues exhibited an increased ability to form sarcospheres. The results of this study provided a significant correlation between resistance and CSCs, and revealed a clue indicating that osteosarcoma recurrence is likely to be associated with CSCs.

This study first systematically analyzed the constituents of an albino mutant strain of Auricularia cornea (AU). After 8 weeks of continuous treatment with metformin (Met) (0.1 g/kg) and AU (0.1 and 0.4 g/kg), db/db mice showed hypoglycemic functioning, indicated by reduced bodyweight, food intake, plasma glucose, serum levels of glycated hemoglobin A1c and glucagon, hepatic levels of phosphoenolpyruvate carboxykinase and lucose-6-phosphatasem, and increased serum levels of insulin. The effect of hypolipidemic functions were indicated by suppressed levels of total cholesterol and triglyceride, and enhanced levels of hepatic glycogen and high-density lipoprotein cholesterol. The renal protective effect of AU was confirmed by the protection in renal structures and the regulation of potential indicators of nephropathy. The anti-oxidative and anti-inflammatory effects of AU were verified by a cytokine array combined with an enzyme-linked immunosorbent assay. AU decreased the expression of protein kinase C α and β2 and phosphor-nuclear factor-κB, and enhanced the expression of catalase, nuclear respiratory factor 2 (Nrf2), manganese superoxide dismutase 2, heme oxygenase-1 and-2, heat shock protein 27 (HSP27), HSP60, and HSP70 in the kidneys of db/db mice. The results confirmed that AU's anti-diabetic and anti-nephritic effects are related to its modulation on oxidative stress.

How to overcome the cell membrane barriers and achieve release payloads efficiently in the cytoplasm have been major challenges for anticancer drug delivery and therapeutic effects with nanosystems. In this study, bovine serum albumin (BSA) was modified with folate acid and histamine, which was then used as the nanocarrier for the antitumor agent doxorubicin (DOX). The DOX-loaded nanoparticles (DOX/FBH-NPs) were prepared via a crosslinking method, and the release of DOX from these nanoparticles (NPs) exhibited pH/reduction-responsive behaviors in vitro. These NPs interacted with the folate receptor overexpressed on the cell membrane of 4 T1 cells and achieved enhanced endocytosis. Afterwards, these NPs exhibited pH-responsiveness within endo-lysosomes and escaped from endosomes due to the "proton sponge" effect, and then completed release of DOX was triggered by high concentration of glutathione (GSH) in cytoplasm. Thus, DOX/FBH-NPs exhibited excellent cytotoxicity against 4 T1 cells in vitro. Benefited from the enhanced permeability and retention (EPR) effect and folate receptor-mediated endocytosis, these NPs gained satisfied tumor-targeting effects in vivo and efficient delivery of DOX to tumor tissues. As a result, these NPs exhibited enhanced antitumor effects and reduced side effects in vivo. In conclusion, these BSA-based NPs modified with both folate acid and histamine showed enhanced tumor-targeting effects in vivo with good biocompatibility and intracellular pH/reduction-responsive behaviors, providing a promising strategy for the efficient delivery of antitumor agents.

Solid nanogenerators often have limited charge transfer due to their low contact area. Liquid-liquid nanogenerators can transfer a charge better than the solid-solid and solid-liquid counterparts. However, the precise manipulation of the liquid morphology remains a challenge because of the fluidity limits of the liquid. In this work, using the surface tension of a droplet to fix its shape, a liquid-liquid triboelectric nanogenerator in Contact-Separation mode is designed using an immiscible aqueous-aqueous interface, achieving a contact surface charge transfer of 129 nC for a single droplet. The configuration is proven to be applicable in humid environments, and the two-phase materials have good biocompatibility and can be used as an effective drug carrier. Therefore, this nanogenerator is useful for designing future implantable devices. Meanwhile, this design also establishes the foundation of aqueous electronics, and additional applications can be achieved using this route.

Oxidative stress triggered by aging, radiation, or inflammation impairs ovarian function by inducing granulosa cell (GC) apoptosis. However, the mechanism inducing GC apoptosis has not been characterized. Here, we found that ovarian GCs from aging patients showed increased oxidative stress, enhanced reactive oxygen species activity, and significantly decreased expression of the known antiapoptotic factor sphingosine-1-phosphate/sphingosine kinase 1 (SPHK1) in GCs. Interestingly, the expression of Krüppel-like factor 12 (KLF12) was significantly increased in the ovarian GCs of aging patients. Furthermore, we determined that KLF12 was significantly upregulated in hydrogen peroxide-treated GCs and a 3-nitropropionic acid-induced in vivo model of ovarian oxidative stress. This phenotype was further confirmed to result from inhibition of SPHK1 by KLF12. Interestingly, when endogenous KLF12 was knocked down, it rescued oxidative stress-induced apoptosis. Meanwhile, supplementation with SPHK1 partially reversed oxidative stress-induced apoptosis. However, this function was lost in SPHK1 with deletion of the binding region to the KLF12 promoter. SPHK1 reversed apoptosis caused by hydrogen peroxide-KLF12 overexpression, a result further confirmed in an in vitro ovarian culture model and an in vivo 3-nitropropionic acid-induced ovarian oxidative stress model. Overall, our study reveals that KLF12 is involved in regulating apoptosis induced by oxidative stress in aging ovarian GCs and that sphingosine-1-phosphate/SPHK1 can rescue GC apoptosis by interacting with KLF12 in negative feedback.

Hepatocellular carcinoma (HCC) is now the second leading cause of cancer death worldwide and lacks effectual therapy due to its high rate of tumor recurrence and metastasis. The aim of this study was to investigate the effects of L antigen family member 3 (LAGE3, a member of the LAGE gene family involved in positive transcription) on the progression of HCC.

High-throughput nanoscopy becomes increasingly important for unraveling complex biological processes from a large heterogeneous cell population at a nanoscale resolution. High-density emitter localization combined with a large field of view and fast imaging frame rate is commonly used to achieve a high imaging throughput, but the image processing speed and the presence of heterogeneous background in the dense emitter scenario remain a bottleneck. Here, we present a simple non-iterative approach, referred to as WindSTORM, to achieve high-speed high-density emitter localization with robust performance for various image characteristics. We demonstrate that WindSTORM improves the computation speed by two orders of magnitude on CPU and three orders of magnitude upon GPU acceleration to realize online image processing, without compromising localization accuracy. Further, WindSTORM is highly robust to maximize the localization accuracy and minimize the image artifacts in the presence of nonuniform background. WindSTORM paves the way for next generation high-throughput nanoscopy.

Single-cell capture plays an important role in single-cell manipulation and analysis. This paper presents a microfluidic device for deterministic single-cell trapping based on the hydrodynamic trapping mechanism. The device is composed of an S-shaped loop channel and thousands of aligned trap units. This arrayed structure enables each row of the device to be treated equally and independently, as it has row periodicity. A theoretical model was established and a simulation was conducted to optimize the key geometric parameters, and the performance was evaluated by conducting experiments on MCF-7 and Jurkat cells. The results showed improvements in single-cell trapping ability, including loading efficiency, capture speed, and the density of the patterned cells. The optimized device can achieve a capture efficiency of up to 100% and single-cell capture efficiency of up to 95%. This device offers 200 trap units in an area of 1 mm², which enables 100 single cells to be observed simultaneously using a microscope with a 20× objective lens. One thousand cells can be trapped sequentially within 2 min; this is faster than the values obtained with previously reported devices. Furthermore, the cells can also be recovered by reversely infusing solutions. The structure can be easily extended to a large scale, and a patterned array with 32,000 trap sites was accomplished on a single chip. This device can be a powerful tool for high-throughput single-cell analysis, cell heterogeneity investigation, and drug screening.

Environmentally persistent organic pollutant (POP) is the general term for refractory organic compounds that show long-range atmospheric transport, environmental persistence, and bioaccumulation. It has been reported that the accumulation of POPs could lead to cellular DNA damage and adverse effects of on metabolic health. To better understand the mechanism of the health risks associated with POPs, we conducted an evidence-based cohort investigation (n = 5,955) at the Jinghai e-waste disposal center in China from 2009 to 2016, where people endure serious POP exposure. And high levels of aging-related diseases, including hypertension, diabetes, autoimmune diseases, and reproductive disorders were identified associated with the POP exposure. In the subsequent molecular level study, an increased telomere dysfunction including telomere multiple telomere signals, telomere signal-free ends, telomere shortening and activation of alternative lengthening of telomeres were observed, which might result from the hypomethylated DNA modification induced telomeric repeat-containing RNA overexpression. Moreover, dysfunctional telomere-leaded senescence-associated secretory phenotype was confirmed, as the proinflammatory cytokines and immunosenescence hallmarks including interleukin-6, P16INK4a, and P14ARF were stimulated. Thus, we proposed that the dysfunctional telomere and elevated systemic chronic inflammation contribute to the aging-associated diseases, which were highly developed among the POP exposure individuals.

The diverse lipid environment of the biological membrane can modulate the structure and function of membrane proteins. However, little is known about the role that lipids play in modulating protein-protein interactions. Here we employed native mass spectrometry (MS) to determine how individual lipid-binding events to the ammonia channel (AmtB) modulate its interaction with the regulatory protein, GlnK. The thermodynamic signature of AmtB-GlnK in the absence of lipids indicates conformational dynamics. A small number of lipids bound to AmtB is sufficient to modulate the interaction with GlnK, and lipids with different headgroups display a range of allosteric modulation. We also find that lipid chain length and stereochemistry can affect the degree of allosteric modulation, indicating an unforeseen selectivity of membrane proteins toward the chemistry of lipid tails. These results demonstrate that individual lipid-binding events can allosterically modulate the interactions of integral membrane and soluble proteins.

The study is aimed to determine the effects of cynarin (Cyn) on mice with gouty arthritis (GA) induced by monosodium urate (MSU). We measured swelling in the hind paws of mice in vivo using Vernier calipers and ultrasound. The liver, kidney, and hind paws were stained with hematoxylin-eosin, and M1 type macrophages were detected in the hind paws using anti-F4/80 and anti-iNOS antibodies. The mRNA expression of inflammatory factors in bone marrow-derived macrophages (BMDMs) and in the hind paws was detected via quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Nucleotide-binding oligomerization domain (NOD)-like receptor (NLR) family pyrin domain containing 3 (NLRP3) inflammasomes and the nuclear factor kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways were analyzed via western blotting. Cyn was detected in vitro using Cell Counting Kit-8 (CCK-8). Cyn treatment reduced hind paw swelling and M1 macrophage infiltration, suppressed the mRNA expression of inflammatory factors, and inhibited NLRP3 inflammasome activation in vivo, in addition to inhibiting the phosphorylation of IKKa/β, p65, and c-Jun NH 2-terminal kinase (JNK). Furthermore, Cyn exerted anti-inflammatory and anti-swelling effects in mice with GA by regulating the NF-κB and JNK pathways and NLRP3 inflammasomes.

A polyethyleneimine (PEI)-assisted copper in-situ growth (CISG) strategy was proposed as a controlled signal amplification strategy to enhance the sensitivity of gold nanoparticle-based lateral flow sensors (AuNP-LFS). The controlled signal amplification is achieved by introducing PEI as a structure-directing agent to regulate the thermodynamics of anisotropic Cu nanoshell growth on the AuNP surface, thus controlling shape and size of the resultant AuNP@Cu core-shell nanostructures and confining free reduction and self-nucleation of Cu2+ for improved reproducibility and decreased false positives. The PEI-CISG-enhanced AuNP-LFS showed ultrahigh sensitivities with the detection limits of 50 fg mL-1 for HIV-1 capsid p24 antigen and 6 CFU mL-1 for Escherichia coli O157:H7. We further demonstrated its clinical diagnostic efficacy by configuring PEI-CISG into a commercial AuNP-LFS detection kit for SARS-CoV-2 antibody detection. Altogether, this work provides a reliable signal amplification platform to dramatically enhance the sensitivity of AuNP-LFS for rapid and accurate diagnostics of various infectious diseases.