Phosphatidylcholine-Plasmalogen-Oleic Acid Reduces BACE1 Expression in Human SH-SY5Y Cells.

Plasmalogens are a family of glycerophospholipids containing one vinyl-ether bond at the sn-1 position in the glycerol backbone, and play important roles in cellular homeostasis including neural transmission. Therefore, reductions of plasmalogens have been associated with neurodegenerative disorders, such as Alzheimer's disease (AD). To evaluate the potential protective effects of plasmalogens against the pathology of AD, protein expression levels of key factors in amyloid precursor protein (APP) metabolic processes were examined using human neuroblastoma SH-SY5Y cells. Here, phosphatidylcholine-plasmalogen-oleic acid (PC-PLS-18) was shown to reduce protein expression levels of ß-site APP cleaving enzyme 1 (BACE1), clusterin, and Tau, factors involved in the amyloid ß-associated pathogenesis of AD. Thus, PC-PLS-18 may have preventive effects against AD by delaying the onset risk for a certain period.

CA9 and PRELID2; hypoxia-responsive potential therapeutic targets for pancreatic ductal adenocarcinoma as per bioinformatics analyses.

A strong hypoxic environment has been observed in pancreatic ductal adenocarcinoma (PDAC) cells, which contributes to drug resistance, tumor progression, and metastasis. Therefore, we performed bioinformatics analyses to investigate potential targets for the treatment of PDAC. To identify potential genes as effective PDAC treatment targets, we selected all genes whose expression level was related to worse overall survival (OS) in The Cancer Genome Atlas (TCGA) database and selected only the genes that matched with the genes upregulated due to hypoxia in pancreatic cancer cells in the dataset obtained from the Gene Expression Omnibus (GEO) database. Although the extracted 107 hypoxia-responsive genes included the genes that were slightly enriched in angiogenic factors, TCGA data analysis revealed that the expression level of endothelial cell (EC) markers did not affect OS. Finally, we selected CA9 and PRELID2 as potential targets for PDAC treatment and elucidated that a CA9 inhibitor, U-104, suppressed pancreatic cancer cell growth more effectively than 5-fluorouracil (5-FU) and PRELID2 siRNA treatment suppressed the cell growth stronger than CA9 siRNA treatment. Thus, we elucidated that specific inhibition of PRELID2 as well as CA9, extracted via exhaustive bioinformatic analyses of clinical datasets, could be a more effective strategy for PDAC treatment.

Individual resolvin E family members work distinctly and in a coordinated manner in the resolution of inflammation.

Three main E-type resolvins (RvEs): RvE1, RvE2, and RvE3, have roles in the resolution of inflammation as anti-inflammatory activities. To investigate the roles of each RvE in the resolution of inflammation, timing of interleukin (IL)- 10 release and IL-10 receptor expressions, and phagocytosis evoked by each RvE in differentiated human monocytes, macrophage-like U937 cells were examined. Here, we show that RvEs enhance the expression of IL-10, and IL-10 receptor-mediated signaling pathways and IL-10-mediated-signaling-independent resolution of inflammatory effects by activating the phagocytotic function. Thus, RvE2 mainly evoked an IL-10-mediated anti-inflammatory function, whereas RvE3 principally activated phagocytotic activity of macrophages, which may be involved in tissue repair. On the other hand, RvE1 showed both functions, although not prominent but rather acting as a relief mediator that takes over the RvE2 function and passes over to the RvE3 function. Therefore, each RvE may act as an important role/stage-specific mediator in a coordinated manner with other RvEs in the processes of the resolution of inflammation.

Pemafibrate inhibited renal dysfunction and fibrosis in a mouse model of adenine-induced chronic kidney disease.

AIMS: Peroxisome proliferator-activated receptor-alpha (PPARα) levels are markedly lower in the kidneys of chronic kidney disease (CKD) patients. Fibrates (PPARα agonists) are therapeutic agents against hypertriglyceridemia and potentially against CKD. However, conventional fibrates are eliminated by renal excretion, limiting their use in patients with impaired renal function. Here, we aimed to evaluate the renal risks associated with conventional fibrates via clinical database analysis and investigate the renoprotective effects of pemafibrate, a novel selective PPARα modulator mainly excreted into the bile. MAIN METHODS: The risks associated with conventional fibrates (fenofibrate, bezafibrate) to the kidneys were evaluated using the Food and Drug Administration Adverse Event Reporting System. Pemafibrate (1 or 0.3 mg/kg/day) was administered daily using an oral sonde. Its renoprotective effects were examined in unilateral ureteral obstruction (UUO)-induced renal fibrosis model mice (UUO mice) and adenine-induced CKD model mice (CKD mice). KEY FINDINGS: The ratios of glomerular filtration rate decreased and blood creatinine increased were markedly higher after conventional fibrate use. Pemafibrate administration suppressed increased gene expressions of collagen-I, fibronectin, and interleukin 1 beta (IL-1ß) in the kidneys of UUO mice. In CKD mice, it suppressed increased plasma creatinine and blood urea nitrogen levels and decreased red blood cell count, hemoglobin, and hematocrit levels, along with renal fibrosis. Moreover, it inhibited the upregulation of monocyte chemoattractant protein-1, IL-1ß, tumor necrosis factor-alpha, and IL-6 in the kidneys of CKD mice. SIGNIFICANCE: These results demonstrated the renoprotective effects of pemafibrate in CKD mice, confirming its potential as a therapeutic agent for renal disorders.

Characterization of Immune Checkpoint Inhibitor-Induced Myasthenia Gravis Using the US Food and Drug Administration Adverse Event Reporting System.

Myasthenia gravis (MG) is a rare but fatal adverse event of immune checkpoint inhibitors (ICIs). We assessed whether patient characteristics differed between those with ICI-related myasthenia gravis and those with idiopathic myasthenia gravis. Reports from the US Food and Drug Administration Adverse Event Reporting System were analyzed. Multivariate analyses were conducted to evaluate the associations between age, sex, and ICI treatment and the reporting rate of myasthenia gravis. Among 5 464 099 cases between 2011 and 2019, 53 447 were treated with ICIs. Myasthenia gravis was reported more often in ICI users. Multiple logistic regression analyses showed that the reporting rate of ICI-related myasthenia gravis did not differ significantly between men and women; however, it was higher in older people than in younger people (adjusted odds ratio, 2.4 [95%CI, 1.84-3.13]). We also investigated useful signs for the early detection of myositis and myocarditis, which are fatal when overlapping with ICI-related myasthenia gravis. Patients with elevated serum creatine kinase or troponin levels were more likely to have concurrent myositis and myocarditis. Unlike idiopathic myasthenia gravis, there was no sex difference in the development of ICI-related myasthenia gravis, which may be more common in older people. Considering the physiological muscle weakness that occurs in the elderly, it may be necessary to monitor ICI-related myasthenia gravis more closely in older people.

Characterization of Human Parainfluenza Virus Receptor Using Terminal Sialic Acid Linkage-Modified Cells.

Human parainfluenza virus type 1 (hPIV1) and type 3 (hPIV3) are respiratory pathogen viruses that bind to terminal sialic acids of glycoconjugates on the cell surface hemagglutinin-neuraminidase glycoprotein. Sialic acid residues are linked to the galactose residue primarily by α2,3 or α2,6 linkages on the terminal of glycoprotein or glycolipids. One of the major determinants of pathogenicity or tissue tropism is virus binding or infection specificity for each sialyl linkage. Sialic linkage-modified human blood cells or mammalian cells that mainly have α2,3- or α2,6-linked sialic acid residues on the surface can be prepared by treatment with linkage-specific sialidases or sialyltransferases. These linkage-modified cells can be used in hemagglutination assays to estimate virus particles' binding specificity, hemadsorption assays to estimate virus glycoproteins' binding specificity, and virus infectivity assays. These methods contribute to identifying the specificity of sialic acid lineage recognition of the hPIV or other sialic acid-binding viruses.

PARP14 regulates EP4 receptor expression in human colon cancer HCA-7 cells.

E type prostanoid 4 (EP4) receptors and their signaling pathways have been implicated in the development and malignant transformation of colorectal cancer. We herein demonstrated that the mono(ADP-ribosyl)ation of histone deacetylase (HDAC)1 and HDAC2 by poly(ADP-ribose) polymerase 14 (PARP14) may be required to induce the expression of EP4 receptors. The suppression of PARP14 activity by siRNA and/or its inhibitors reduced the mRNA expression of EP4 receptors. Thus, the expression of their proteins to approximately 50-80% in human colon cancer HCA-7 cells, however, which retained the activities of EP4 receptors to some extent. Since the expression levels of EP4 receptors are important factors for the maintenance of homeostasis, the adequate inhibition of PARP14 activity will be a good target for the prevention of colon cancer and/or as an alternative therapy for this disease. Since non-steroidal anti-inflammatory drugs (NSAIDs) are associated with a risk of heart attacks and stroke, novel PARP14 inhibitors will supersede NSAIDs without causing heart attacks and stroke, while maintaining appropriate EP4 receptor-mediated intestinal homeostasis.

Identification and Characterization of Human Colorectal Cancer Cluster Predominantly Expressing EP3 Prostanoid Receptor Subtype.

Colorectal cancer (CRC) is one of the common types of cancer in humans. Prostaglandin E2 (PGE2) is a well-known mediator of colorectal cancer through stimulation of four E-type prostanoid (EP) receptor subtypes: EP1, EP2, EP3, and EP4 receptors. All subtypes of EP receptors are involved in CRC promotion or malignancy. However, the characteristics of CRC that highly expresses EP receptor subtypes have not been clarified. In the present study, we classified CRC from a cancer genomic database and identified CRC clusters which highly express EP receptor subtypes. Most of these clusters predominantly expressed one subtype of EP receptor and showed different gene expression patterns. Among them, we focused on the cluster highly expressing the EP3 receptor (CL-EP3). As the result of characterization of gene expression, CL-EP3 was characterized as: epithelial mesenchymal transition (EMT)-induced progressed cancer with activation of transforming growth factor-ß pathway, activation of hypoxia-inducible factor-1α, and suppression of runt-related transcription factor 3. Since we previously reported that EP3 receptor is involved in and induce colon cancer cell migration, EP3 receptor-expressing CRC may induce metastasis through these signaling pathways. Thus, the findings suggest the effectiveness of cancer clustering by gene expression of the EP receptor subtype to elucidate the mechanism of human CRC.

Down-regulation of the expression of cyclooxygenase-2 and prostaglandin E2 by interleukin-4 is mediated via a reduction in the expression of prostanoid EP4 receptors in HCA-7 human colon cancer cells.

Chronic inflammatory bowel disease (IBD), which is characterized by prolonged inflammation of the gastrointestinal tract is associated with an increased risk of colorectal cancer. Recent studies revealed that the pathology of IBD is caused by hyperactivated immune responses mediated by differentiated CD4+ naïve helper T cells, such as Th1 and Th17 cells, but not Th2 cells. The human E-type prostanoid 4 (EP4) receptor and its pathways have also been implicated in and/or associated with the early developmental stages of colorectal cancer along with increases in the levels of prostaglandin E2 (PGE2) and cyclooxygenase-2 (COX-2), the hallmarks of colorectal carcinogenesis. In the present study, using an in silico analysis and pharmacological experiments, we demonstrated that interleukin (IL)-4, a signature cytokine of Th2 cells, down-regulated the expression of COX-2 and PGE2 in the human colon cancer cell line, HCA-7. This result may be attributed to a reduction in the expression of prostanoid EP4 receptors through the induction of hypoxia inducible factor-1α via the interleukin-4 receptor-stimulated activation of signal transducer and activator of transcription 6. However, another major Th2 cytokine IL-13 had no effect on the expression of COX-2 or prostanoid EP4 receptors in HCA-7 cells. Therefore, instead of the hyperactivation of Th1/Th17 cells, the deactivation/down-regulation of Th2 cells followed by a decrease in the production of IL-4 in IBD may play a role in the cancerous transformation of cells, at least in prostanoid EP4 receptor-overactivated tumorigenesis.

Identification of prophylactic drugs for oxaliplatin-induced peripheral neuropathy using big data.

BACKGROUND: Drug repositioning is a cost-effective method to identify novel disease indications for approved drugs; it requires a shorter developmental period than conventional drug discovery methods. We aimed to identify prophylactic drugs for oxaliplatin-induced peripheral neuropathy by drug repositioning using data from large-scale medical information and life science information databases. METHODS: Herein, we analyzed the reported data between 2007 and 2017 retrieved from the FDA's database of spontaneous adverse event reports (FAERS) and the LINCS database provided by the National Institute of Health. The efficacy of the drug candidates for oxaliplatin-induced peripheral neuropathy obtained from the database analysis was examined using a rat model of peripheral neuropathy. Additionally, we compared the incidence of peripheral neuropathy in patients who received oxaliplatin at the Tokushima University Hospital, Japan. The effects of statins on the animal model were examined in six-week-old male Sprague-Dawley rats and seven or eight-week-old male BALB/C mice. Retrospective medical chart review included clinical data from Tokushima University Hospital from April 2009 to March 2018. RESULTS: Simvastatin, indicated for dyslipidemia, significantly reduced the severity of peripheral neuropathy and oxaliplatin-induced hyperalgesia. In the nerve tissue of model rats, the mRNA expression of Gstm1 increased with statin administration. A retrospective medical chart review using clinical data revealed that the incidence of peripheral neuropathy decreased with statin use. CONCLUSION AND RELEVANCE: Thus, drug repositioning using data from large-scale basic and clinical databases enables the discovery of new indications for approved drugs with a high probability of success.

Phosphatidylcholine-Plasmalogen-Oleic Acid Has Protective Effects against Arachidonic Acid-Induced Cytotoxicity.

Plasmalogens are a group of glycerophospholipids containing a vinyl-ether bond at the sn-1 position in the glycerol backbone. Cellular membrane plasmalogens are considered to have important roles in homeostasis as endogenous antioxidants, differentiation, and intracellular signal transduction pathways including neural transmission. Therefore, reduced levels of plasmalogens have been suggested to be associated with neurodegenerative diseases such as Alzheimer's disease. Interestingly, although arachidonic acid is considered to be involved in learning and memory, it could be liberated and excessively activate neuronal activity to the excitotoxic levels seen in Alzheimer's disease patients. Here, we examined the protective effects of several kinds of plasmalogens against cellular toxicity caused by arachidonic acid in human neuroblastoma SH-SY5Y cells. As a result, only phosphatidylcholine-plasmalogen-oleic acid (PC-PLS-18) showed protective effects against arachidonic acid-induced cytotoxicity based on the results of lactate dehydrogenase release and ATP depletion assays, as well as cellular morphological changes in SH-SY5Y cells. These results indicate that PC-PLS-18 protects against arachidonic acid-induced cytotoxicity, possibly via improving the stability of the cellular membrane in SH-SY5Y cells.

The Gαs-protein-mediated pathway may be steadily stimulated by prostanoid EP2 receptors, but not by EP4 receptors.

EP2 and EP4 prostanoid receptors have long been considered to have similar roles, since they are known to couple with Gαs-protein and activate cAMP-mediated signaling pathways. In this study, we re-evaluated the results of cAMP assays with or without phosphodiesterase (PDE) inhibitor pretreatment. Here, we show that in the absence of PDE inhibitor pretreatment, prostaglandin E2 causes accumulation of cAMP in EP2 receptors, whereas markedly low levels of cAMP accumulated in EP4 receptors. By applying the Black/Leff operational model calculation, we found that EP2 receptors have a biased ability to intrinsically activate the Gαs-protein-mediated pathway, whereas EP4 receptors have strong biased activity for the Gαi-protein-mediated pathway. Thus, EP2 and EP4 receptors may not be similar Gαs-coupled receptors but instead substantially different receptors with distinct roles.

The differential functional coupling of phosphodiesterase 4 to human DP and EP2 prostanoid receptors stimulated with PGD2 or PGE2.

BACKGROUND: Human DP and EP2 receptors are two of the most homologically related receptors coupling with Gαs-protein, which stimulate adenylyl cyclase to produce cAMP. Indeed, both receptors are considered to be generated by tandem duplication. It has been reported that other highly homologous and closely related ß1- and ß2-adrenergic receptors interact distinctly with and differentially regulate cAMP-specific phosphodiesterase (PDE) 4 recruitment. METHODS: First, we focused on the cAMP degradation pathways of DP and EP2 receptors stimulated by prostaglandin (PG) D2 or PGE2 using HEK cells stably expressing either human DP receptors or EP2 receptors. Then, distances between ligands and amino acids of the receptors were evaluated by molecular dynamics (MD) analysis. RESULTS: We found that PGD2/EP2 receptors exerted a greater effect on PDE4 activity than PGE2/EP2 receptors. Moreover, by MD analysis, either the PGD2 or EP2 receptor was moved and the distance was shortened between them. According to the results, DP receptors retain reactivity for PGE2, but EP2 receptors may be activated only by PGE2, at least in terms of cAMP formation, through the differential functional coupling of PDE4 probably with ß-arrestin. CONCLUSION: Since DP receptors and EP2 receptors are considered to be duplicated genes, DP receptors may still be in a rapid evolutionary stage as a duplicated copy of EP2 receptors and have not yet sufficient selectivity for their cognate ligand, PGD2.

Development of a mugineic acid family phytosiderophore analog as an iron fertilizer.

Iron (Fe) is an essential nutrient, but is poorly bioavailable because of its low solubility in alkaline soils; this leads to reduced agricultural productivity. To overcome this problem, we first showed that the soil application of synthetic 2'-deoxymugineic acid, a natural phytosiderophore from the Poaceae, can recover Fe deficiency in rice grown in calcareous soil. However, the high cost and poor stability of synthetic 2'-deoxymugineic acid preclude its agricultural use. In this work, we develop a more stable and less expensive analog, proline-2'-deoxymugineic acid, and demonstrate its practical synthesis and transport of its Fe-chelated form across the plasma membrane by Fe(III)•2'-deoxymugineic acid transporters. Possibility of its use as an iron fertilizer on alkaline soils is supported by promotion of rice growth in a calcareous soil by soil application of metal free proline-2'-deoxymugineic acid.

Diphenhydramine may be a preventive medicine against cisplatin-induced kidney toxicity.

Cisplatin is widely used as an anti-tumor drug for the treatment of solid tumors. Unfortunately, it causes kidney toxicity as a critical side effect, limiting its use, given that no preventive drug against cisplatin-induced kidney toxicity is currently available. Here, based on a repositioning analysis of the Food and Drug Administration Adverse Events Reporting System, we found that a previously developed drug, diphenhydramine, may provide a novel treatment for cisplatin-induced kidney toxicity. To confirm this, the actual efficacy of diphenhydramine was evaluated in in vitro and in vivo experiments. Diphenhydramine inhibited cisplatin-induced cell death in kidney proximal tubular cells. Mice administered cisplatin developed kidney injury with significant dysfunction (mean plasma creatinine: 0.43 vs 0.15 mg/dl) and showed augmented oxidative stress, increased apoptosis, elevated inflammatory cytokines, and MAPKs activation. However, most of these symptoms were suppressed by treatment with diphenhydramine. Furthermore, the concentration of cisplatin in the kidney was significantly attenuated in diphenhydramine-treated mice (mean platinum content: 70.0 vs 53.4 µg/g dry kidney weight). Importantly, diphenhydramine did not influence or interfere with the anti-tumor effect of cisplatin in any of the in vitro or in vivo experiments. In a selected cohort of 98 1:1 matched patients from a retrospective database of 1467 patients showed that patients with malignant cancer who had used diphenhydramine before cisplatin treatment exhibited significantly less acute kidney injury compared to ones who did not (6.1 % vs 22.4 %, respectively). Thus, diphenhydramine demonstrated efficacy as a novel preventive medicine against cisplatin-induced kidney toxicity.

15-Keto-PGE2 acts as a biased/partial agonist to terminate PGE2-evoked signaling.

Prostaglandin E2 (PGE2) is well-known as an endogenous proinflammatory prostanoid synthesized from arachidonic acid by the activation of cyclooxygenase-2. E type prostanoid (EP) receptors are cognates for PGE2 that have four main subtypes: EP1 to EP4. Of these, the EP2 and EP4 prostanoid receptors have been shown to couple to Gαs-protein and can activate adenylyl cyclase to form cAMP. Studies suggest that EP4 receptors are involved in colorectal homeostasis and cancer development, but further work is needed to identify the roles of EP2 receptors in these functions. After sufficient inflammation has been evoked by PGE2, it is metabolized to 15-keto-PGE2 Thus, 15-keto-PGE2 has long been considered an inactive metabolite of PGE2 However, it may have an additional role as a biased and/or partial agonist capable of taking over the actions of PGE2 to gradually terminate reactions. Here, using cell-based experiments and in silico simulations, we show that PGE2-activated EP4 receptor-mediated signaling may evoke the primary initiating reaction of the cells, which would take over the 15-keto-PGE2-activated EP2 receptor-mediated signaling after PGE2 is metabolized to 15-keto-PGE2 The present results shed light on new aspects of 15-keto-PGE2, which may have important roles in passing on activities to EP2 receptors from PGE2-stimulated EP4 receptors as a "switched agonist." This novel mechanism may be significant for gradually terminating PGE2-evoked inflammation and/or maintaining homeostasis of colorectal tissues/cells functions.

Fibroblast-specific ERK5 deficiency changes tumor vasculature and exacerbates tumor progression in a mouse model.

The roles of cancer-associated fibroblasts (CAFs) have been studied in the tumor progression, and CAFs are expected to become the new targets for cancer pharmacotherapies. CAFs contribute to tumor cell survival and proliferation, tumor angiogenesis, immune suppression, tumor inflammation, tumor cell invasion and metastasis, and extracellular matrix remodeling. However, detailed mechanisms of how CAFs function in the living system remain unclear. CAFs include α-smooth muscle actin, expressing activated fibroblasts similar to myofibroblasts, and are highly capable of producing collagen. Several reports have demonstrated the contributions of extracellular-signal-regulated kinase 5 (ERK5) in fibroblasts to the fibrotic processes; however, the roles of CAF-derived ERK5 remain unclear. To investigate the roles of CAF-derived ERK5 in the tumor progression, we created mice lacking the ERK5 gene specifically in fibroblasts. Colon-26 mouse colon cancer cells were implanted into the mice subcutaneously, and the histological analyses of the tumor tissue were performed after 2 weeks. Immunofluorescence analyses showed that recipient-derived fibroblasts existed within the tumor tissue. The present study demonstrated that fibroblast-specific ERK5 deficiency exacerbated tumor progression and it was accompanied with thicker tumor vessel formation and the increase in the number of activated fibroblasts. We combined the results of The Cancer Genome Atlas (TCGA) database analysis with our animal studies, and indicated that regulating ERK5 activity in CAFs or CAF invasion into the tumor tissue can be important strategies for the development of new targets in cancer pharmacotherapies.

Proton pump inhibitors block iron absorption through direct regulation of hepcidin via the aryl hydrocarbon receptor-mediated pathway.

Proton pump inhibitors (PPIs) have been used worldwide to treat gastrointestinal disorders. A recent study showed that long-term use of PPIs caused iron deficiency; however, it is unclear whether PPIs affect iron metabolism directly. We investigated the effect of PPIs on the peptide hepcidin, an important iron regulatory hormone. First, we used the FDA Adverse Event Reporting System database and analyzed the influence of PPIs. We found that PPIs, as well as H2 blockers, increased the odds ratio of iron-deficient anemia. Next, HepG2 cells were used to examine the action of PPIs and H2 blockers on hepcidin. PPIs augmented hepcidin expression, while H2 blockers did not. In fact, the PPI omeprazole increased hepcidin secretion, and omeprazole-induced hepcidin upregulation was inhibited by gene silencing or the pharmacological inhibition of the aryl hydrocarbon receptor. In mouse experiments, omeprazole also increased hepatic hepcidin mRNA expression and blood hepcidin levels. In mice treated with omeprazole, protein levels of duodenal and splenic ferroportin decreased. Taken together, PPIs directly affect iron metabolism by suppressing iron absorption through the inhibition of duodenal ferroportin via hepcidin upregulation. These findings provide a new insight into the molecular mechanism of PPI-induced iron deficiency.

A I131V Substitution in the Fusion Glycoprotein of Human Parainfluenza Virus Type 1 Enhances Syncytium Formation and Virus Growth.

Human parainfluenza virus type 1 (hPIV1) has two spike glycoproteins, the hemagglutinin-neuraminidase (HN) glycoprotein as a receptor-binding protein and the fusion (F) glycoprotein as a membrane-fusion protein. The F glycoprotein mediates both membrane fusion between the virus and cell and membrane fusion between cells, called syncytium formation. Wild-type C35 strain (WT) of hPIV1 shows little syncytium formation of infected cells during virus growth. In the present study, we isolated a variant virus (Vr) from the WT that showed enhanced syncytium formation of infected cells by using our previously established hPIV1 plaque formation assay. Vr formed a larger focus and showed increased virus growth compared with WT. Sequence analysis of the spike glycoprotein genes showed that the Vr had a single amino acid substitution of Ile to Val at position 131 in the fusion peptide region of the F glycoprotein without any substitutions of the HN glycoprotein. The Vr F glycoprotein showed enhanced syncytium formation in F and HN glycoprotein-expressing cells. Additionally, expression of the Vr F glycoprotein increased the focus area of the WT-infected cells. The single amino acid substitution at position 131 in the F glycoprotein of hPIV1 gives hPIV1 abilities to enhance syncytium formation and increase cell-to-cell spread. The present study supports the possibility that hPIV1 acquires increased virus growth in vitro from promotion of direct cell-to-cell transmission by syncytium formation.