The minimally important difference (MID) refers to the smallest change that is sufficiently meaningful to carry implications for patients' care. MIDs are necessary to guide the interpretation of scores. This study estimated MID for the Patient Reported Outcomes Measurement Information System (PROMIS) pain interference (PI).

The identification of cancer stem-like cells is a recent development in ovarian cancer. Compared to other cancer cells, cancer stem-like cells present more chemo-resistance and more aggressive characteristics. They play an important role in the recurrence and drug resistance of cancer. Therefore, the target therapy of cancer stem-like cell may become a promising and effective approach for ovarian cancer treatment. It may also help to provide novel diagnostic and therapeutic strategies.

SYP-1018 is a lyophilized polymeric nanoparticle formulation of voriconazole that is under development for intravenous dosing. This study compared the pharmacokinetic and tolerability profiles of SYP-1018 with those of Vfend(®), the marketed formulation of voriconazole. The effect of CYP2C19 polymorphism on the voriconazole pharmacokinetics was also evaluated.

This study proposes a tactile estimation method of molded plastic plates based on human tactile perception characteristics. Plastic plates are often used in consumer products. The tactile evaluation plays an important role in product development. However, physical quantities not taking into account human tactile perception have been employed in previous tactile estimation procedures. Hence, in this study, we adopted the vibrational thresholds of the mechanoreceptive units-FA I, FA II, SA I and SA II-for stimuli detection and developed a tactile estimation method for plastic plates that clarified the mechanoreceptive units related to tactile sensation. The developed tactile sensor consists of a base and a silicone rubber pad that contains strain gauges in it. We detected vibration during touch by the sensor and calculated the estimation of the firing values of the cutaneous mechanoreceptors, which are the essential data obtained by humans during tactile perception, in comparison to the amplitude spectrum of the vibration with the threshold amplitude of each mechanoreceptive unit. Simultaneously, we calculated the relationship between the normal and tangential forces recorded while the sensor ran over the samples. As a result of stepwise linear regression analysis using these values as explanatory variables, the evaluation scores for Soft were successfully estimated using the firing value of FA II and the relationship between normal/tangential forces, and the evaluation scores for Rough were estimated using the SA I firing value.

Activation of the Wnt/β-catenin pathway plays a pathogenic role in age-related macular degeneration (AMD) and is thus a potential target for the development of therapeutics for this disease. Here, we demonstrated that Wnt5a antagonized β-catenin response transcription (CRT) induced with Wnt3a by promoting β-catenin phosphorylation at Ser33/Ser37/Thr41 and its subsequent degradation in human retinal pigment epithelial (RPE) cells. Wnt5a decreased the levels of vascular endothelial growth factor (VEGF), tumor necrosis factor-α (TNF-α), and nuclear factor-κB (NF-κB), which was up-regulated by Wnt3a. Furthermore, Wnt5a increased E-cadherin expression and decreased cell migration by down-regulating Snail expression, thereby abrogating the Wnt3a-induced epithelial-mesenchymal transition (EMT) in human RPE cells. Our findings suggest that Wnt5a suppresses the pathogenic effects of canonical Wnt signaling in human RPE cells by promoting β-catenin phosphorylation and degradation. Therefore, Wnt5a has significant therapeutic potential for the treatment of AMD.

Cell detachment is essential in culturing adherent cells. Trypsinization is the most popular detachment technique, even though it reduces viability due to the damage to the membrane and extracellular matrix. Avoiding such damage would improve cell culture efficiency. Here we propose an enzyme-free cell detachment method that employs the acoustic pressure, sloshing in serum-free medium from intermittent traveling wave. This method detaches 96.2% of the cells, and increases its transfer yield to 130% of conventional methods for 48 h, compared to the number of cells detached by trypsinization. We show the elimination of trypsinization reduces cell damage, improving the survival of the detached cells. Acoustic pressure applied to the cells and media sloshing from the intermittent traveling wave were identified as the most important factors leading to cell detachment. This proposed method will improve biopharmaceutical production by expediting the amplification of tissue-cultured cells through a more efficient transfer process.

We investigated the characteristics of microvessel tortuosity in branch retinal vein occlusion (BRVO) and central retinal vein occlusion (CRVO) and their associations with visual outcomes using optical coherence tomography angiography (OCTA). Thirty-four BRVO and 21 CRVO patients and 31 healthy subjects were included. From OCTA, the branch number (BN), mean branch length (BL), mean Euclidean length (EL), vessel density (VD) and vessel tortuosity (VT) were quantified. In BRVO eyes, compared with that in the controls, the affected area of the deep capillary plexus (DCP) showed a decreased BN and VD, an increased BL, and unchanged VT. The nonaffected area of the DCP showed decreases in BN, VD and VT. The affected area of the superficial capillary plexus (SCP) showed higher VT. In CRVO eyes, the DCP showed a lower BN, VD and VT, while the SCP showed a lower BN and greater BL and EL. Improved visual acuity (VA) after 1 year in BRVO eyes was associated with decreases in BN, BL, VD and VT in the affected area in the DCP and lower VT in the nonaffected area of the SCP; in CRVO eyes, improved VA was associated with a higher BL and EL in the DCP. VT, BL, and EL may be new microvascular markers associated with changes in VA in BRVO and CRVO.

Constant or intense light degenerates the retina and retinal pigment epithelial cells. Light generates reactive oxygen species and nitric oxide leading to initial reactions of retinal degeneration. Apoptosis is the primary mechanism of abnormal death of photoreceptors, retinal ganglion cells, or retinal pigment epithelium (RPE) in degenerative retinal diseases, including diabetic retinopathy and age-related macular degeneration. The current study evaluated the function of erythropoietin (EPO) on angiogenesis and apoptosis in the retina and RPE under oxidative stress. We determined the pro-angiogenic and antiapoptotic mechanism of EPO under stress conditions using a conditional EPO knockdown model using siRNA, EPO addition, proteomics, immunocytochemistry, and bioinformatic analysis. Our studies verified that EPO protected retinal cells from light-, hypoxia-, hyperoxia-, and hydrogen peroxide-induced apoptosis through caspase inhibition, whereas up-regulated angiogenic reactions through vascular endothelial growth factor (VEGF) and angiotensin pathway. We demonstrated that the EPO expression in the retina and subsequent serine/threonine/tyrosine kinase phosphorylations might be linked to oxidative stress response tightly to determining angiogenesis and apoptosis. Neuroprotective roles of EPO may involve the balance between antiapoptotic and pro-angiogenic signaling molecules, including BCL-xL, c-FOS, caspase-3, nitric oxide, angiotensin, and VEGF receptor. Our data indicate a new therapeutic application of EPO toward retinal degeneration based on the dual roles in apoptosis and angiogenesis at the molecular level under oxidative stress.

Neovascular age-related macular degeneration (nAMD) is a common cause of irreversible vision loss in the elderly. Anti-vascular endothelial growth factor has been effective in treating pathological ocular neovascularization, but it has limitations including the need for repeated intraocular injections for the maintenance of therapeutic effects in most patients and poor or non-response to this agent in some patients. in vitro cellular studies were conducted using retinal pigment epithelial cell lines (ARPE-19 and hTERT-RPE1), human umbilical vein endothelial cells (HUVECs), and human umbilical vein smooth muscle cells (HUVSMCs). in vivo efficacy of ilimaquinone (IQ) was tested in laser-induced choroidal neovascularization mouse and rabbit models. Tissue distribution study was performed in male C57BL6/J mice. IQ, 4,9-friedodrimane-type sesquiterpenoid isolated from the marine sponge, repressed the expression of angiogenic/inflammatory factors and restored the expression of E-cadherin in retinal pigment epithelial cells by inhibiting the Wnt/β-catenin pathway. In addition, it selectively inhibited proliferation and tube formation of HUVECs by activating the p53 pathway. Topical and intraperitoneal administration of IQ significantly reduced choroidal neovascularization in rabbits and mice with laser-induced choroidal neovascularization. Notably, IQ by the oral route of exposure was highly permeable to the eyes and suppressed abnormal vascular leakage by downregulation of β-catenin and stabilization of p53 in vivo. Our findings demonstrate that IQ functions through regulation of p53 and Wnt/β-catenin pathways with conceivable advantages over existing cytokine-targeted anti-angiogenic therapies.

We investigated the association of visual outcome in typical neovascular age-related macular degeneration (nAMD) and polypoidal choroidal vasculopathy (PCV) with or without pachychoroid with lesion areas on optical coherence tomography (OCT) quantified by convolutional neural network (CNN) analysis. Treatment-naïve 132 nAMD and 45 PCV eyes treated with ranibizumab or aflibercept for at least 12 months were retrospectively reviewed. Significant factors, including intraretinal fluid (IRF), subretinal fluid (SRF), pigment epithelial detachment (PED) and subretinal hyperreflective material (SHRM) area quantified by CNN at baseline and 12 months, were analyzed by logistic regression analyses for 3-line visual gain or maintenance of 20/30 Snellen vision. Visual gain at the final visit in nAMD was associated with a smaller SHRM at baseline (OR 0.167, P = 0.03), greater decrease in SRF and SHRM from baseline to month 12 (OR 1.564, P = 0.02; OR 12.877, P = 0.01, respectively). Visual gain in nAMD without pachychoroid was associated with a greater decrease in SRF and SHRM (OR 1.574, P = 0.03, OR 1.775, P = 0.04). No association was found in nAMD with pachychoroid and any type of PCV. Greater decrease in SRF and SHRM from baseline to month 12 was associated with favorable visual outcomes in nAMD without pachychoroid but not in nAMD with pachychoroid and PCV.

The development of molecular imaging probes to identify key cellular changes within lung metastases may lead to noninvasive detection of metastatic lesions in the lung. In this study, we constructed a macrophage-targeted clickable albumin nanoplatform (CAN) decorated with mannose as the targeting ligand using a click reaction to maintain the intrinsic properties of albumin in vivo. We also modified the number of mannose molecules on the CAN and found that mannosylated serum albumin (MSA) harboring six molecules of mannose displayed favorable pharmacokinetics that allowed high-contrast imaging of the lung, rendering it suitable for in vivo visualization of lung metastases. Due to the optimized control of functionalization and surface modification, MSA enhanced blood circulation time and active/passive targeting abilities and was specifically incorporated by mannose receptor (CD206)-expressing macrophages in the metastatic lung. Moreover, extensive in vivo imaging studies using single-photon emission computed tomography (SPECT)/CT and positron emission tomography (PET) revealed that blood circulation of time-optimized MSA can be used to discern metastatic lesions, with a strong correlation between its signal and metastatic burden in the lung.

Three-dimensional cell agglomerates are broadly useful in tissue engineering and drug testing. We report a well-free method to form large (1.4-mm) multicellular clusters using 100-MHz surface acoustic waves (SAW) without direct contact with the media or cells. A fluid couplant is used to transform the SAW into acoustic streaming in the cell-laden media held in a petri dish. The couplant transmits longitudinal sound waves, forming a Lamb wave in the petri dish that, in turn, produces longitudinal sound in the media. Due to recirculation, human embryonic kidney (HEK293) cells in the dish are carried to the center of the coupling location, forming a cluster in less than 10 min. A few minutes later, these clusters may then be translated and merged to form large agglomerations, and even repeatedly folded to produce a roughly spherical shape of over 1.4 mm in diameter for incubation-without damaging the existing intercellular bonds. Calcium ion signaling through these clusters and confocal images of multiprotein junctional complexes suggest a continuous tissue construct: intercellular communication. They may be formed at will, and the method is feasibly useful for formation of numerous agglomerates in a single petri dish.

Background: In recent years, there has been considerable interest in the therapeutic targeting of tumor-associated macrophages (TAMs) to modulate the tumor microenvironment (TME), resulting in antitumoral phenotypes. However, key mediators suitable for TAM-mediated remodeling of the TME remain poorly understood. Methods: In this study, we used single-cell RNA sequencing analyses to analyze the landscape of the TME modulated by TAMs in terms of a protumor microenvironment during early tumor development. Results: Our data revealed that the depletion of TAMs leads to a decreased epithelial-to-mesenchymal transition (EMT) signature in cancer cells and a distinct transcriptional state characterized by CD8+ T cell activation. Moreover, notable alterations in gene expression were observed upon the depletion of TAMs, identifying Galectin-1 (Gal-1) as a crucial molecular factor responsible for the observed effect. Gal-1 inhibition reversed immune suppression via the reinvigoration of CD8+ T cells, impairing tumor growth and potentiating immune checkpoint inhibitors in breast tumor models. Conclusion: These results provide comprehensive insights into TAM-mediated early tumor microenvironments and reveal immune evasion mechanisms that can be targeted by Gal-1 to induce antitumor immune responses.

Age-related macular degeneration (AMD) is the leading cause of blindness in the world. Evidence indicates that the suppression of the ubiquitin-proteasome system (UPS) contributes to the accumulation of toxic proteins and inflammation in retinal pigment epithelium (RPE), the functional abnormalities and/or the degeneration of which are believed to be the initiators and major pathologies of AMD. To identify new protein associations with the altered UPS in AMD, we used LC-ESI-MS/MS to perform a proteomic analysis of the aqueous humor (AH) of AMD patients and matched control subjects. Six UPS-related proteins were present in the AH of the patients and control subjects. Four of the proteins, including 26S proteasome non-ATPase regulatory subunit 1 (Rpn2), were increased in patients, according to semi-quantitative proteomic profiling. An LC-MRM assay revealed a significant increase of Rpn2 in 15 AMD patients compared to the control subjects, suggesting that this protein could be a biomarker for AMD.

Protein phosphorylations have essential regulatory roles in visual signaling. Previously, we found that phosphorylation of several proteins in the retina and retinal pigment epithelium (RPE) is involved in anti-apoptotic signaling under oxidative stress conditions, including light exposure. In this study, we used a phosphoprotein enrichment strategy to evaluate the light-induced phosphoproteome of primary bovine RPE cells. Phosphoprotein-enriched extracts from bovine RPE cells exposed to light or dark conditions for 1h were separated by 2D SDS-PAGE. Serine and tyrosine phosphorylations were visualized by 2D phospho Western blotting and specific phosphorylation sites were analyzed by tandem mass spectrometry. Light induced a marked increase in tyrosine phosphorylation of beta crystallin A3 and A4. The most abundant light-induced up-regulated phosphoproteins were crystallins of 15-25 kDa, including beta crystallin S and zeta crystallin. Phosphorylation of beta crystallin suggests an anti-apoptotic chaperone function of crystallins in the RPE. Other chaperones, cytoskeletal proteins, and proteins involved in energy balance were expressed at higher levels in the dark. A detailed analysis of RPE phosphoproteins provides a molecular basis for understanding of light-induced signal transduction and anti-apoptosis mechanisms. Our data indicates that phosphorylation of crystallins likely represents an important mechanism for RPE shielding from physiological and pathophysiological light-induced oxidative injury.

Albumin is one of the most attractive nanoplatforms for targeted imaging and drug delivery due to its biocompatibility and long circulation half-life. However, previously reported albumin-based nanoplatforms have shown inconsistent blood circulation half-life according to the modified methods, and the affecting factors were not well evaluated, which could hamper the clinical translation of albumin-based nanoplatforms. Herein, we developed a finely tuned click-chemistry based albumin nanoplatform (CAN) with a longer circulation half-life and an efficient tumor targeting ability. Methods: CAN was synthesized in two steps. First, albumin was conjugated with ADIBO-NHS (albumin-ADIBO) by reacting albumin with various molar ratios of ADIBO. The number of attached ADIBO moieties was determined using matrix-assisted laser desorption ionization time of flight (MALDI-TOF). Second, the desired modalities including azide-functionalized chelator, a fluorescence dye, and folate were incorporated into albumin-ADIBO using strain-promoted alkyne-azide cycloaddition reaction (SPAAC reaction). The biodistribution and targeting efficiency of functionalized CANs were demonstrated in mice. Results: The degree of functionalization (DOF) and resulting in vivo biodistribution was controlled precisely using the click chemistry approach. Specifically, the numbers of attached azadibenzocyclooctyne (ADIBO) moieties on albumin, the DOF, were optimized by reacting albumin with varying molar ratios of ADIBO with a high reproducibility. Furthermore, we developed a simple and efficient method to estimate the DOF using UV-visible spectrophotometry (UV-vis), which was further validated by matrix-assisted laser desorption ionization time of flight (MALDI-TOF). The biodistribution of CAN could be controlled by DOF, and CAN with an optimized DOF showed a long circulation half-life (> 18 h). CAN was further functionalized using a simple click chemistry reaction with an azide functionalized chelator, a fluorescence dye, and folate. 64Cu- and folate-labeled CAN (64Cu-CAN-FA) showed effective and specific folate receptor targeting in vivo, with an over two-fold higher uptake than the liver at 24 h post-injection. Conclusions: Our development from the precisely controlled DOF demonstrates that an optimized CAN can be used as a multifunctional nanoplatform to obtain a longer half-life with radioisotopes and ligands, and provides an effective method for the development of albumin-based tumor theranostic agents.

As current therapies benefit only a minority of cancer patients, additional therapeutic targets are needed. Tumor-associated macrophages (TAMs) have attracted attention for improving therapeutic responses, yet regulatory strategies remain elusive. Here, we show that the protein kinase A catalytic subunit (PKA-C) acts as a molecular switch, inducing a pro-tumoral immunosuppressive macrophage phenotype within tumors. In human and murine breast cancer, overactivated PKA in TAMs creates a detrimental microenvironment for cancer progression by inducing vascular endothelial growth factor A (VEGFA), interleukin-10 (IL-10), and macrophage-derived arginase 1 (ARG1) expression. Macrophages with genetic deletion of PKA-C are prone to be pro-inflammatory, suggesting a possible immunotherapeutic target. Delivery of liposomal PKA inhibitor facilitates tumor regression and abrogates pro-tumoral TAM functions in mice. The therapeutic effect of targeting PKA is pronounced when combined with αCTLA-4 antibody, increasing cluster of differentiation 8 (CD8)+GranzymeB+ T cells by about 60-fold. Our findings demonstrate critical roles of TAM PKA-C in tumor progression and suggest that targeting PKA-C efficiently augments cancer treatment responses.

To generate three-dimensional tissue in vitro, promoting vasculogenesis in cell aggregates is an important factor. Here, we found that ultrasound promoted vasculogenesis of human umbilical vein endothelial cells (HUVECs). Promotion of HUVEC network formation and lumen formation were observed using our method. In addition to morphological evaluations, protein expression was quantified by western blot assays. As a result, expression of proteins related to vasculogenesis and the response to mechanical stress on cells was enhanced by exposure to ultrasound. Although several previous studies have shown that ultrasound may promote vasculogenesis, the effect of ultrasound was unclear because of unregulated ultrasound, the complex culture environment, or two-dimensional-cultured HUVECs that cannot form a lumen structure. In this study, regulated ultrasound was propagated on three-dimensional-monocultured HUVECs, which clarified the effect of ultrasound on vasculogenesis. We believe this finding may be an innovation in the tissue engineering field.

Hyperthermia has been studied as a noninvasive cancer treatment. Cancer cells show stronger thermal cytotoxicity than normal cells, which is exploited in hyperthermia. However, the absence of methods evaluating the thermal cytotoxicity in cells prevents the development of hyperthermia. To investigate the thermal cytotoxicity, culture temperature should be regulated. We, thus, developed a culture system regulating culture temperature immediately and accurately by employing metallic culture vessels. Michigan Cancer Foundation-7 cells and normal human dermal fibroblasts were used for models of cancer and normal cells. The findings showed cancer cells showed stronger thermal cytotoxicity than normal cells, which is quantitatively different from previous reports. This difference might be due to regulated culture temperature. The thermal stimulus condition (43 °C/30 min) was, further, focused for assays. The mRNA expression involving apoptosis changed dramatically in cancer cells, indicating the strong apoptotic trend. In contrast, the mRNA expression of heat shock protein (HSP) of normal cells upon the thermal stimulus was stronger than cancer cells. Furthermore, exclusively in normal cells, HSP localization to nucleus was confirmed. These movement of HSP confer thermotolerance to cells, which is consistent with the different thermal cytotoxicity between cancer and normal cells. In summary, our developed system can be used to develop hyperthermia treatment.

Metabolic dysregulation is an important hallmark of cancer. Nicotinamide (NAM), a water-soluble amide form of niacin (vitamin B3), is currently available as a supplement for maintaining general physiologic functions. NAM is a crucial regulator of mitochondrial metabolism and redox reactions. In this study, we aimed to identify the mechanistic link between NAM-induced metabolic regulation and the therapeutic efficacy of NAM in triple-negative breast cancer (TNBC). The combined analysis using multiomics systems biology showed that NAM decreased mitochondrial membrane potential and ATP production, but increased the activities of reverse electron transport (RET), fatty acid β-oxidation and glycerophospholipid/sphingolipid metabolic pathways in TNBC, collectively leading to an increase in the levels of reactive oxygen species (ROS). The increased ROS levels triggered apoptosis and suppressed tumour growth and metastasis of TNBC in both human organoids and xenograft mouse models. Our results showed that NAM treatment leads to cancer cell death in TNBC via mitochondrial dysfunction and activation of ROS by bifurcating metabolic pathways (RET and lipid metabolism); this provides insights into the repositioning of NAM supplement as a next-generation anti-metabolic agent for TNBC treatment.