Cellular Ca²⁺ signaling functions as one of the most common second messengers of various signal transduction pathways in cells and mediates a number of physiological roles in a cell-type dependent manner. Ca²⁺ signaling also regulates more general and fundamental cellular activities, including cell proliferation and apoptosis. Among ion channels, Ca²⁺-permeable channels in the plasma membrane as well as endo- and sarcoplasmic reticulum membranes play important roles in Ca²⁺ signaling by directly contributing to the influx of Ca²⁺ from extracellular spaces or its release from storage sites, respectively. Furthermore, Ca²⁺-gated ion channels in the plasma membrane often crosstalk reciprocally with Ca²⁺ signals and are central to the regulation of cellular functions. This review focuses on the physiological and pharmacological impact of i) Ca²⁺-gated ion channels as an apparatus for the conversion of cellular Ca²⁺ signals to intercellularly propagative electrical signals and ii) the opposite feedback regulation of Ca²⁺ signaling by Ca²⁺-gated ion channel activities in excitable and non-excitable cells.

With the development of structural biology and data mining, computer-aided drug design (CADD) has been playing an important role in all aspects of new drug development. Reverse docking, a method of virtual screening based on molecular docking in CADD, is widely used in drug repositioning, drug rescue, and traditional Chinese medicine (TCM) research, for it can search for macromolecular targets that can bind to a given ligand molecule. This review revealed the principle of reverse docking, summarized common target protein databases and docking procedures, and enumerated the applications of reverse docking in drug repositioning, adverse drug reactions, traditional Chinese medicine, and coronavirus disease 2019 (COVID-19) treatment. Hope our work can give some inspiration to researchers engaged in drug development.

This study aimed to explore the effect of curcumin and hydromorphone hydrochloride (HH) cotreatment on postoperative pain in rats. An incision + formaldehyde-induced pain rat model was established. Rats were treated with vehicle, curcumin, HH, or curcumin + HH. Paw mechanical withdrawal threshold and thermal withdrawal latency were measured at 1 d before surgery as well as 1 , 2 h, 1 , 3 , and 7 d after surgery to assess pain sensitivity. The L4-6 region of the spinal cord was collected from each rat at 2 h, 1 , 3 , and 7 d after surgery. Western blot analysis and immunohistochemical staining were carried out to detect the protein expression of pain-related genes. Quantitative real-time PCR and enzyme-linked immunosorbent assay were conducted to measure the expression and production of proinflammatory mediators. Compared with other groups, Curcumin + HH significantly reduced pain sensitivity in the model rats. Mechanistically, curcumin + HH suppressed protein expression of stromal cell-derived factor-1 (SDF-1), CXC chemokine receptor 4 (CXCR4), p-Akt, and c-fos while enhancing protein expression of nerve growth factor (NGF) in the dorsal root ganglia (DRG) of model rats. Curcumin + HH inhibited the expression and production of interleukin 1β (IL-1β), cyclooxygenase-2 (COX-2), tumor necrosis factor α (TNF-α), and p65 nuclear factor kappa B (NF-κB) in the DRG. Coadministration of curcumin and HH alleviates incision + formaldehyde-induced pain in rats, possibly by suppressing the SDF-1/CXCR4 pathway and the production of proinflammatory mediators. Our results provide curcumin and HH cotreatment as a promising therapeutic strategy in the management of postoperative pain.

Imatinib mesylate is a potent tyrosine kinase inhibitor that may induce immunological effects, such as inhibition of immune suppressive cells; but, how it modulates the immune system remains to be completely elucidated. In this study, we showed that cell proliferation of CT26 colon cancer and Lewis lung carcinoma (3LL) lung cancer cells was not inhibited by imatinib in vitro, although its administration significantly suppressed the growth of CT26, but not 3LL, subcutaneous tumors, and prolonged survival in CT26 tumor-bearing mice. Further, we examined the expression of immune cell-related molecules in the tumors to elucidate the differences in imatinib-mediated antitumor effects between CT26 and 3LL tumors. The nCounter assay showed that the expression of CD8 and CD8⁺ T cell-recruiting chemokine genes was significantly elevated in imatinib-treated CT26 tumors than that in control tumors; however, the gene expression remained unchanged in imatinib-treated or control 3LL tumors. Furthermore, frequency of interferon-γ⁺ (IFN-γ⁺) CD8⁺ T cells was increased in imatinib-treated CT26 tumors than control tumors, indicating induction of antitumor immunity by imatinib. The analysis indicates that imatinib promotes infiltration of effector T cells in tumors by upregulating expression of cytokines that recruit CD8⁺ T cells in the tumor microenvironment, which may lead to a strong antitumor effect.

We aimed to evaluate the efficacy and safety of propranolol gel at various concentrations with infantile hemangiomas after proliferative phases. We designed a single-center, randomized, double-blind, dose-dependent trial with placebo control and randomized patients to receive propranolol gel at 0, 1, or 5%, twice daily for 24 weeks. The primary efficacy endpoint was the percentage change in redness of the tumors. Safety endpoints were skin characteristics changes and systemic symptoms. We made two comparisons to evaluate the superiority of 1 and 5% propranolol gels against placebo for primary endpoint analysis and used the t-test to compare parents’ satisfaction with these treatments. Initially, 19 patients were enrolled, but 8 were excluded from the analysis. We were underpowered to answer the question of efficacy. In the per-protocol set, we found similar results for the redness percentage change among the patients on placebo, 1 and 5% gel. However, the difference in redness before and after treatment suggested a slight decreasing trend of lesion’s redness as the propranolol concentration increased. The difference in parents’ satisfaction between the placebo and 5% propranolol gel groups was significant (p = 0.08). We observed no serious adverse events. We did not find an obvious dose-dependent effect for the propranolol gel treatment against infantile hemangiomas after the proliferative phase. However, external applications twice daily were less burdensome for parents and led to good compliances. It had a favorable safety profile in Japanese pediatric patients with infantile hemangiomas.

Methylglyoxal (MGO), which is produced as a byproduct of glucose metabolism, is the leading to diabetic cardiovascular complications. Salvia miltiorrhiza Bunge (Lamiaceae) has been reported as a potential plant to control diabetes and cardiovascular disease. However, no report exists on the effect of Salvia miltiorrhiza Bunge extract (SME) on MGO-induced glucotoxicity in human umbilical vein endothelial cells (HUVECs). We demonstrated the protective effects of SME (1, 5, and 10 µg/mL) and its components against MGO-induced endothelial dysfunction in HUVECs. Cytotoxicity was evaluated using the several in vitro experiments. Additionally, the protein expression of receptor of advanced glycation end-products (RAGE), mitogen-activated protein kinase (MAPK) pathway and glyoxalase system were measured. Then, the inhibitory effects of SME and its main components on MGO-induced oxidative stress, radical scavenging, formation of MGO-derived advanced glycation end products (AGEs), and MGO-AGEs crosslinking were evaluated. SME (10 µg/mL) strongly prevented expressed levels of RAGE, MGO-induced apoptosis and reduced reactive oxygen species (ROS) generation in HUVECs, comparing with 1 mM aminoguanidine. Additionally, SME (5 and 10 µg/mL) reduced the expression of proteins (e.g., p-extracellular signal-regulated kinase (ERK) and p-p38) in the MAPKs pathway and upregulated the glyoxalase system in HUVECs. SME (0.5–10 mg/mL), dihydrotanshinone (0.4 mM), and rosmarinic acid (0.4 mM) prevented MGO-AGEs formation and broke the MGO-AGE crosslinking. These results show that S. miltiorrhiza has protective effects against MGO-induced glucotoxicity by regulating the proteins involved in apoptosis, glyoxalase system and antioxidant activity. We expect that S. miltiorrhiza is a potential natural resource for the treatment of MGO-induced vascular endothelial dysfunction.

Gambogenic acid (GNA) is extracted from plant Gamboge, has a wide range of anti-tumor effects. In this paper, we study the inhibitory effect of GNA on the BEL-7402/ADM of hepatoma resistant cell lines and further study the mechanism of action. Cell viability test represented that GNA could improve the sensitivity of hepatoma drug-resistant cell line BEL-7402/ADM to Adriamycin (ADM), and further study by 4′,6-diamidino-2-phenylindole (DAPI) staining and flow cytometry found that GNA could improve the effect of ADM on promoting apoptosis in BEL-7402/ADM cells, and the activation of apoptosis-related protein was significantly increased, and the ratio of Bax to Bcl-2 was significantly increased. Monodansylcadaverine staining and transmission electron microscopy showed that the basal autophagy level of BEL-7402/ADM cells was higher than that of BEL-7402 cells. Further detection of protein expression found that the intracellular LC3-II to LC3-I ratio and Beclin 1 protein expression increased in the combination of GNA and ADM, but the protein level of p62 increased significantly. GNA inhibit protective autophagy in BEL-7402/ADM cells and promote the role of ADM in inducing apoptosis, thereby increasing ADM sensitivity to BEL-7402/ADM cells, and the effect of GNA inhibition of autophagy may be achieved by inhibiting the degradation of autophagosomes.

The ginsenoside Rbs are the primary active compounds of Panax ginseng and ginsenoside Rb2 is a renowned component among the Rbs. This study aimed to investigate the potential effects of ginsenoside Rb2 on coronary heart disease (CHD). H9c2 cells were exposed to H₂O₂ to establish CHD model in vitro. Gene expression was determined by quantitative realtime PCR (qPCR) and Western blot. Cellular functions were detected by Cell Counting Kit-8 (CCK-8), 5-ethynyl-2′-deoxyuridine (EdU), flow cytometry, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) assays. We found that Ginsenoside Rb2 promoted cell proliferation while suppressed oxidative stress and apoptosis of H9c2 cells induced by H₂O₂ exposure. Mechanistically, Ginsenodise Rb2 involves in the regulation of nuclear factor, erythroid 2 like 2 (Nrf2)/heme oxygenase (HO)-1 signaling pathway. Inactivation of Nrf2/HO-1 signaling pathway reversed the effects of ginsenoside Rb2 on H9c2 cells. Taken together, ginsenoside Rb2 exhibited a cardioprotective effect in vitro. The underlying mechanism of ginsenoside Rb2 in H9c2 cells could be standardized to Nrf2/HO-1 signaling pathway, inhibiting cell apoptosis and regaining cell proliferation. The present study has proposed a novel mechanism of ginsenoside Rb2 in the cardioprotective effect.

Cyclooxygenase (COX)-1-selective inhibitors have side effects such as itching and dryness of the skin. In this study, the degree of skin dryness and the onset mechanism of this condition were investigated by comparing the effects of three non-steroidal anti-inflammatory drugs (NSAIDs) in mice. Mice were orally administered either indomethacin, loxoprofen sodium, or celecoxib (n = 5 per group) once daily for four consecutive days, and blood samples as well as skin and jejunal tissues were isolated on day 5. In the mice treated with indomethacin, transepidermal water loss was significantly increased, and dry skin was observed. In addition, the expression of matrix metalloproteinase (MMP)-I, mast cells, CD163, CD23, CD21, histamine, and peroxisome proliferation-activated receptor (PPAR)γ in the skin and jejunum was increased, and the blood levels of interleukin-10 and immunoglobulin E were also increased. In contrast, the expression of collagen type I in the skin was decreased. These results show that indomethacin activates PPARγ in the skin and jejunum, changes the polarity of macrophages, increases the secretion of MMP-1 from mast cells, and decomposes collagen type I, leading to dry skin.

Status epilepticus (SE) induced inflammation plays an important role in the pathogenesis of SE. Long non-coding RNA small nucleolar RNA host gene 5 (lncRNA Snhg5) has been reported in various inflammatory diseases. However, the mechanism of Snhg5 regulated inflammation in SE remains unclear. Therefore, this study aimed to clarify the role and mechanism of Snhg5 in SE-induced inflammation in vitro and vivo. In vitro, lipopolysaccharide (LPS)-induced inflammation in microglia was used to mimic the inflammation after SE. In vivo, SE model was induced by lithium chloride and pilocarpine. The level of Snhg5, p65, p-p65, p-inhibitor of kappaB (IκB)α, IκBα and inflammatory factors (tumor necrosis factor (TNF)-α, interleukin (IL)-1β) were measured via quantitative real-time PCR or Western blot. The Nissl stain and immunohistochemical stain were performed to observe hippocampal damage and microglia proliferation. The results showed Snhg5 was up-regulated in the rat and microglia. Knockdown of Snhg5 inhibited LPS-induced inflammation and relative expression of p-65/p65, p-IκBα/IκBα. Moreover, down-regulation of Snhg5 attenuated SE-induced inflammation and reduced the number of microglia in hippocampus. These findings indicated that Snhg5 modulates the inflammation via nuclear factor-kappaB (NF-κB) signaling pathway in SE rats.

Our previous studies identified that nimesulide analogs which bear a methoxy substituent at the para-position of the phenyl ring could be potential radiotracer candidates for detecting disorders related to cyclooxygenase-2 (COX-2) expression and activity in vivo using positron emission tomography (PET) in the brain. The present study was conducted to evaluate the in vivo characteristics of ¹¹C-labeled para-methoxy nimesulide ([¹¹C]1d) as a brain COX-2-targeted imaging agent compared to other isomeric methoxy analogs of nimesulide ([¹¹C]1b and [¹¹C]1c). [¹¹C]1b–d were synthesized with reasonable yield and purity by the methylation of the O-desmethyl precursor with [¹¹C]methyl triflate in the presence of NaOH at room temperature. We performed in vivo biodistribution analysis, brain PET imaging, ex vivo autoradiography, and metabolite analysis in mice. The uptake of [¹¹C]1b–d was lower in the brain than in other tissues, including in the blood, and both [¹¹C]1c and [¹¹C]1d were rapidly metabolized. However, [¹¹C]1d showed a small, but significant, specific signal and heterogeneous distribution in the brain. In vivo evaluation suggested that [¹¹C]1d might correlate with COX-2 expression in the brain. Given its instability in vivo, [¹¹C]1d seems unsuitable as a brain-COX-2 radioimaging agent. Further structural refinement of these radiotracers is necessary to enhance their uptake in the brain and to achieve sufficient metabolic stability.

Individual differences in gut microbiota can affect the pharmacokinetics of drugs. Yokukansan is a traditional Japanese kampo medicine used to treat peripheral symptoms of dementia and delirium. A study examining the pharmacokinetics of the components of yokukansan reported large individual differences in the pharmacokinetics of glycyrrhizic acid (GL). It is known that GL is metabolized by intestinal bacteria to glycyrrhetinic acid (GA), which is absorbed in the gastrointestinal tract. Thus, the gut microbiota may affect GL pharmacokinetics. We aimed to clarify the relationship between the gut microbiota composition and pharmacokinetics of GL in yokukansan. Mice were orally administered yokukansan, following the administration of various antibiotics, and the plasma concentration of GA and composition of gut microbiota were measured. The GA plasma concentration was low in mice treated with amoxicillin and vancomycin. The composition of gut microbiota revealed a different pattern from that of the control group. Mice with low plasma levels of GA had lower levels of the phylum Bacteroides and Firmicutes. Additionally, bacteria, such as those belonging to the genera Parabaceroides, Bacteroides, Ruminococcus and an unknown genus in families Lachnospiraceae and Ruminococcaceae, exerted positive correlations between the gene copies and plasma GA levels. These bacteria may contribute to the absorption of GA in the gastrointestinal tract, and multiple bacteria may be involved in GL pharmacokinetics. The pharmacokinetics of GL may be predicted by evaluating the composition of gut bacteria, rather than by evaluating the amount of a single bacterium.

Dysgeusia is a major side effect of anti-cancer drug treatment. Since dysgeusia significantly lowers the patient’s QOL, predicting and avoiding its onset in advance is desirable. Accordingly, aims of the present study were to use a genome-wide association study (GWAS) to identify genes associated with the development of dysgeusia in patients taking anti-cancer drugs and to predict the development of dysgeusia using associated single nucleotide polymorphisms (SNPs). GWAS was conducted on 76 patients admitted to the Department of Hematology, Tokushima University Hospital. Using Sanger sequencing for 23 separately collected validation samples, the top two SNPs associated with the development of dysgeusia were determined. GWAS identified rs73049478 and rs41396146 SNPs on the retinoic acid receptor beta (RARB) gene associated with dysgeusia development due to the administration of anti-cancer drugs. Evaluation of the two SNPs using 23 validation samples indicated that the accuracy rate of rs73049478 was relatively high (87.0%). Thus, the findings of the present study suggest that the rs73049478 SNP of RARB can be used to predict the onset of dysgeusia caused by the administration of anti-cancer drugs.

Medication wastage is a global issue; however, there are few reports in Japanese hospitals. The purpose of this study was to clarify the situation of medication wastage at our university hospital. We investigated the numbers, costs, reasons, occurrence departments, and involvement of high-priced medications of medication wastage for two years. We analyzed 6730 cases and the total cost was 22782027 Japanese yen (JPY). The most common reasons for medication wastage were change or discontinuation of medication after preparation and breakage or contamination due to dropping. The highest cost was expired medications. The department with the highest number of cases was the hospital wards; however, the hospital pharmacy department accounted for the majority of the costs and most of the reasons were expired medications. Medication wastage of 50000 JPY or more per case was only 1.3% of the total but accounted for 70.7% of the cost and medication wastage of 100000 JPY or more per case was only 0.7% of the total but accounted for 58.6% of the cost. These findings indicate that expired medications in the hospital pharmacy department have the largest impact on medication wastage from the viewpoint of economic loss, and suggest the need for efforts on medication management focusing on high-priced medications. The challenge of minimizing medication wastage should be addressed from the perspectives of both hospital management and the effective use of resources.

Physiologically based pharmacokinetic (PBPK) modeling has the potential to play significant roles in estimating internal chemical exposures. The three major PBPK model input parameters (i.e., absorption rate constants, volumes of the systemic circulation, and hepatic intrinsic clearances) were generated in silico for 212 chemicals using machine learning algorithms. These input parameters were calculated based on sets of between 17 and 65 chemical properties that were generated by in silico prediction tools before being processed by machine learning algorithms. The resulting simplified PBPK models were used to estimate plasma concentrations after virtual oral administrations in humans. The estimated absorption rate constants, volumes of the systemic circulation, and hepatic intrinsic clearance values for the 212 test compounds determined traditionally (i.e., based on fitting to measured concentration profiles) and newly estimated had correlation coefficients of 0.65, 0.68, and 0.77 (p < 0.01, n = 212), respectively. When human plasma concentrations were modeled using traditionally determined input parameters and again using in silico estimated input parameters, the two sets of maximum plasma concentrations (r = 0.85, p < 0.01, n = 212) and areas under the curve (r = 0.80, p < 0.01, n = 212) were correlated. Virtual chemical exposure levels in liver and kidney were also estimated using these simplified PBPK models along with human plasma levels. These results indicate that the PBPK model input parameters for humans of a diverse set of compounds can be reliability estimated using chemical descriptors calculated using in silico tools.

The purpose of this study was to develop a Bio-layer interferometry (BLI) system that could be an alternative approach for the direct evaluation of anti-polyethylene glycol (PEG) immunoglobulin M (IgM)-mediated complement activation of the accelerated blood clearance (ABC) phenomenon. Complement activation is well known to play an important role in the clearance of PEGylated and non-PEGylated nanomedicines following intravenous injection. This complement system is also thought to be responsible for the ABC phenomenon wherein repeated injections of PEGylated products are bound by anti-PEG antibodies. This study used three different sources of anti-PEG antibodies: HIK-M09 monoclonal antibodies (mAbs); HIK-M11 mAbs; and antiserum containing polyclonal anti-PEG IgMs. 1,2-Distearoyl-sn-glycero-3-phosphoethanolamine-n-[methoxy (polyethylene glycol)-2000] (mPEG₂₀₀₀-DSPE) was immobilized as an antigen on aminopropyl silane biosensor chips of BLI. All anti-PEG IgMs in the sources increased the signals (thickness of the layer around the sensor tip) regarding binding of anti-PEG antibodies to PEG on the chips. In all anti-PEG IgM sources, further increases in the signals were observed when incubated in naïve mouse serum, which is a complement source, but not in heat inactivated (56 °C, 30 min) mouse serum, which abolishes complement activity. These findings show that the complement activation mediated via anti-PEG IgMs, which occurred on the sensor chips, was detected via BLI analysis. The complement activation induced by all anti-PEG IgM sources was confirmed via conventional enzyme-linked immunosorbent assay (ELISA), which is the conventional mode for detection of complement activation. Our study results show that BLI is a simple alternative method for the detection of complement activation.

Warfarin is a representative anticoagulant with large interindividual variability. The published kinetic-pharmacodynamic (K-PD) model allows the prediction of warfarin dose requirement in Swedish patients; however, its applicability in Japanese patients is not known. We evaluated the model’s predictive performance in Japanese patients with various backgrounds and relationships using Bayesian parameter estimation and sampling times. A single-center retrospective observational study was conducted at Tokyo Women’s Medical University, Medical Center East. The study population consisted of adult patients aged >20 years who commenced warfarin with a prothrombin time-international normalized ratio (PT-INR) from June 2015 to June 2019. The published K-PD model modified by Wright and Duffull was assessed using prediction-corrected visual predictive checks, focusing on clinical characteristics, including age, renal function, and individual prediction error. The external dataset included 232 patients who received an initial warfarin daily dose of 3.2 ± 1.28 mg with 2278 PT-INR points (median [range] follow-up period of 23 d [7–28]). Prediction-corrected visual predictive checks carried a propensity for underprediction. Additionally, age >60 years, body mass index ≤25 kg/m², and estimated glomerular filtration rate ≤60 mL/min/1.73 m² had a pronounced tendency to underpredict PT-INR. However, Bayesian prediction using four prior observations reduced underprediction. To improve the prediction performance of these special populations, further studies are required to construct a model to predict warfarin dose requirements in Japanese patients.

To investigate the mechanism of the protective effect of tetrahydroxystilbene glucoside (TSG) on nerve cells, an injury model induced by rotenone in PC12 cells was constructed. Cell viability was detected by using cell counting kit-8 (CCK8) assay. Apoptosis was detected by using flow cytometry. The mitochondrial membrane potential (MMP) was detected by using the fluorescent probe JC-1. Generation of reactive oxygen species (ROS) in PC12 cells was determined using the 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate acetyl ester (CM-H₂DCFDA) probe. Protein expression in PC12 cells was detected using Western blotting. The results showed that TSG (20–100 µM) attenuated the cytotoxic effects of rotenone on PC12 cells. TSG pretreatment attenuated the apoptosis rate, the degradation of poly(ADP-ribose)polymerase (PARP) and the activation of cleaved caspase 3, which was induced by rotenone. TSG can significantly reduce the effect of rotenone on the reduction of MMP and the expression of cytoC in the cytosolic fraction. TSG attenuated rotenone-induced de-phosphorylation and mitochondrial translocation of cofilin, as well as rotenone-induced accumulation of ROS. The Western blot results showed that ROT could decrease the expression level of phosphorylated (p)-Glycogen synthase kinase-3β (GSK)-3β and p-AKT, and TSG could weaken these effects of rotenone. In addition, TSG increased the expression level of nuclear factor-E2-related factor 2 (Nrf2) in the nuclear fraction. These results suggest that TSG could protect PC12 cells against rotenone through multiple pathways. Thus, TSG has the potential to become a novel neuroprotective agent.

The aim of this work is to develop a new assay system for screening biliary excretion drugs. When monolayers of human liver-derived cell lines HepG2 and Huh-7 were grown on an insert membrane, the efflux ratio (ER: ratio of the apparent permeability coefficient in the basal-to-apical direction (P_app,B-to-A) to that in the apical to basal direction (P_app,A-to-B)) of sulfobromophthalein (BSP), a model substrate of multidrug resistance-associated protein 2 (MRP2), was greater than 1.0, indicating transport of BSP in the efflux direction. The efflux transport was significantly suppressed by MK-571, an inhibitor of MRPs, in both cell lines. Expression of MRP2 mRNA in HepG2 and Huh-7 was 3.5- and 1.4-fold higher, respectively, than in primary human hepatocytes, while expression of P-glycoprotein and breast cancer resistance protein mRNAs was markedly lower, supporting the idea that MRP2 is the main mediator of directional BSP transport in this assay system. The advantage of our system is the potential to quantitatively evaluate biliary excretion of MRP2 substrates in vitro.