Glycolytic activity is required for the onset of neural plate folding during neural tube closure in mouse embryos.

Physiological hypoxia is critical for placental mammalian development. However, the underlying mechanisms by which hypoxia regulates embryonic development remain unclear. We discovered that the expression of glycolytic genes partially depends on hypoxia in neuroepithelial cells of E8.25 mouse embryos. Consistent with this finding, inhibiting glycolysis during the early phase of neural tube closure (E8.0-8.5) resulted in a neural tube closure defect. In contrast, inhibiting the electron transport chain did not affect neural tube formation. Furthermore, inhibiting glycolysis affected cell proliferation, but not differentiation and survival. Inhibiting glycolysis repressed the phosphorylation of myosin light chain 2, and consequent neural plate folding. Our findings revealed that anaerobic glycolysis regulates neuroepithelial cell proliferation and apical constriction during the early phase of neural tube closure.

LIF-IGF Axis Contributes to the Proliferation of Neural Progenitor Cells in Developing Rat Cerebrum.

In rodent models, leukemia inhibitory factor (LIF) is involved in cerebral development via the placenta, and maternal immune activation is linked to psychiatric disorders in the child. However, whether LIF acts directly on neural progenitor cells (NPCs) remains unclear. This study performed DNA microarray analysis and quantitative RT-PCR on the fetal cerebrum after maternal intraperitoneal or fetal intracerebral ventricular injection of LIF at day 14.5 (E14.5) and determined that the expression of insulin-like growth factors (IGF)-1 and -2 was induced by LIF. Physiological IGF-1 and IGF-2 levels in fetal cerebrospinal fluid (CSF) increased from E15.5 to E17.5, following the physiological surge of LIF levels in CSF at E15.5. Immunostaining showed that IGF-1 was expressed in the cerebrum at E15.5 to E19.5 and IGF-2 at E15.5 to E17.5 and that IGF-1 receptor and insulin receptor were co-expressed in NPCs. Further, LIF treatment enhanced cultured NPC proliferation, which was reduced by picropodophyllin, an IGF-1 receptor inhibitor, even under LIF supplementation. Our findings suggest that IGF expression and release from the NPCs of the fetal cerebrum in fetal CSF is induced by LIF, thus supporting the involvement of the LIF-IGF axis in cerebral cortical development in an autocrine/paracrine manner.

Hif1α-dependent hypoxia signaling contributes to the survival of deep-layer neurons and cortex formation in a mouse model.

Hypoxia-inducible factor 1 α (Hif1α) plays a crucial role in brain development. To study the function of Hif1α in early brain development, we generated neuroepithelial cell-specific Hif1α-knockout mice. Hif1α-knockout mice died soon after birth; these mice exhibited an abnormal head shape, indicating the presence of brain defects. Morphological analysis revealed that Hif1α ablation reduced the overall size of the brain, especially affecting the telencephalon. Neuronal apoptosis predominantly occurred in deep-layer neurons, consequently the alignment of cortical layers was severely disorganized in Hif1α knockout mice. Furthermore, we demonstrated that Vegf signaling contributes to the survival of deep-layer neurons as a downstream effector of Hif1α-dependent hypoxia signaling. Taken together, our findings demonstrate that Hif1α plays a critical role in the early stages of telencephalon development.

Altered gut microbiota and its association with inflammation in patients with chronic thromboembolic pulmonary hypertension: a single-center observational study in Japan.

BACKGROUND: The pathogenesis of chronic thromboembolic pulmonary hypertension (CTEPH) is considered to be associated with chronic inflammation; however, the underlying mechanism remains unclear. Recently, altered gut microbiota were found in patients with pulmonary arterial hypertension (PAH) and in experimental PAH models. The aim of this study was to characterize the gut microbiota in patients with CTEPH and assess the relationship between gut dysbiosis and inflammation in CTEPH. METHODS: In this observational study, fecal samples were collected from 11 patients with CTEPH and 22 healthy participants. The abundance of gut microbiota in these fecal samples was assessed using 16S ribosomal ribonucleic acid (rRNA) gene sequencing. Inflammatory cytokine and endotoxin levels were also assessed in patients with CTEPH and control participants. RESULTS: The levels of serum tumor necrosis factor-α (TNF-α), interleukin (IL)-6, IL-8, and macrophage inflammatory protein (MIP)-1α were elevated in patients with CTEPH. Plasma endotoxin levels were significantly increased in patients with CTEPH (P < 0.001), and were positively correlated with TNF-α, IL-6, IL-8, and MIP-1α levels. The 16S rRNA gene sequencing and the principal coordinate analysis revealed the distinction in the gut microbiota between patients with CTEPH (P < 0.01) and control participants as well as the decreased bacterial alpha-diversity in patients with CTEPH. A random forest analysis for predicting the distinction in gut microbiota revealed an accuracy of 80.3%. CONCLUSION: The composition of the gut microbiota in patients with CTEPH was distinct from that of healthy participants, which may be associated with the elevated inflammatory cytokines and endotoxins in CTEPH.

Crystal structure and conformational stability of a galectin-1 tandem-repeat mutant with a short linker.

Modification of the domain architecture of galectins has been attempted to analyze their biological functions and to develop medical applications. Several types of galectin-1 repeat mutants were previously reported but, however, it was not clear whether the native structure of the wild type was retained. In this study, we determined the crystal structure of a galectin-1 tandem-repeat mutant with a short linker peptide, and compared the unfolding profiles of the wild type and mutant by chemical denaturation. The structure of the mutant was consistent with that of the dimer of the wild type, and both carbohydrate-binding sites were retained. The unfolding curve of the wild type with lactose suggested that the dimer dissociation and the tertiary structure unfolding was concomitant at micromolar protein concentrations. The midpoint denaturant concentration of the wild type was dependent on the protein concentration and lower than that of the mutant. Linking the two subunits significantly stabilized the tertiary structure. The mutant exhibited higher T-cell growth-inhibition activity and comparable hemagglutinating activity. Structural stabilization may prevent the oxidation of the internal cysteine residue.

MC5R Contributes to Sensitivity to UVB Waves and Barrier Function in Mouse Epidermis.

MC5R is known for its role in the exocrine function of sebaceous glands, but other functions in the epidermis remain unclear. This study focused on the relationship between MC5R and homeostasis in the epidermis and examined the role of MC5R in mice whose skin was irradiated with UVB waves. UVB irradiation-induced skin ulcers and severe inflammation at lower doses in homozygotes of MC5R-deficient (i.e., MC5R -/- ) mice (150 mJ/cm2) than the doses in wild-type mice (500 mJ/cm2). Transepidermal water loss was increased (approximately 10-fold) in adult MC5R -/- mice compared with that in wild-type mice. In neonates, a dye exclusion assay showed no remarkable difference between MC5R -/- and wild-type mice. After UVB irradiation, compared with wild-type mice, MC5R -/- mice showed increased inflammatory cell infiltration in the dermis of the ulcerative region, significantly increased thickness of the epidermis in the nonulcerative region, significantly more prickle cells in the nonulcerative region, and increased serum IL-6 levels but decreased IL-10 levels. Transmission electron microscopy revealed fewer lamellar granules, less lipid secretion, and an expansion of the trans-Golgi network in the epidermis in MC5R -/- mice. This study elucidated the increased sensitivity to UVB irradiation and decreased barrier function in MC5R -/- mice.

Leukemia Inhibitory Factor Induces Proopiomelanocortin via CRH/CRHR Pathway in Mouse Trophoblast.

We previously showed that maternal leukemia inhibitory factor (LIF) induces placental production of adrenocorticotropic hormone (ACTH), which stimulates fetal nucleated red blood cells to further secrete LIF and promote neurogenesis in rodent brains. However, the underlying mechanism of LIF-dependent ACTH induction remains unclear. Recently, we found that LIF induces corticotropin-releasing hormone (CRH) in mouse trophoblast stem cells. This finding supports the results of a previous study that CRH, which is produced by the placenta, induces placental ACTH production. In this study, we examined whether the effects of LIF are mediated by the induction of Pomc via CRH upregulation in mouse trophoblast. In vivo, protein levels of LIF and CRH peak in mouse placenta at 13.5 days post coitum. In mouse placenta, Crh mRNA and protein levels significantly increased 3 h after intraperitoneal injection of LIF (5 µg/kg body weight) into dams at 13.5 days post coitum. We also examined the effect of LIF-induced CRH on the expression of Pomc induced by LIF in mouse trophoblast stem cells in vitro. After LIF supplementation for 3 days, we found that the increased expression of Crh-induced by new supplementation of LIF was earlier than that of Pomc. Furthermore, LIF-induced upregulation of Pomc in mouse trophoblast stem cells was attenuated by inhibition of the CRH/CRHR1 pathway, whereas LIF-induced secretion of ACTH was attenuated by inhibition of the JAK/STAT3 pathway. Therefore, LIF indirectly increases placental Pomc expression through the CRH/CRHR1 pathway, and placental ACTH secretion is induced directly by LIF via the JAK/STAT3 pathway.

Mid-pregnancy maternal immune activation increases Pax6-positive and Tbr2-positive neural progenitor cells and causes integrated stress response in the fetal brain in a mouse model of maternal viral infection.

Maternal immune activation (MIA) in midpregnancy is a risk factor for neurodevelopmental disorders. Improper brain development may cause malformations of the brain; maldevelopment induced by MIA may lead to a pathology-related phenotype. In this study, a single intraperitoneal injection of 20 mg/kg polyriboinosinic-polyribocytidylic acid [poly(I:C)] was administered to C57BL/6J mice on embryonic day (E) 12.5 to mimic maternal viral infection. Histopathological analysis of neurogenesis was performed using markers for Pax6, Tbr2, and Tbr1. In these fetuses, significant increases were observed in the proportion of Pax6-positive neural progenitor cells and Pax6/Tbr2 double-positive cells 24 h after poly(I:C) injection. There were no differences in the proportion of Tbr1-positive postmitotic neurons 48 h after poly(I:C) injection. At E18.5, there were more Pax6-positive and Tbr2-positive neural progenitor cells in the poly(I:C)-injected group than in the saline-injected group. Gene ontology enrichment analysis of poly(I:C)-induced differentially expressed genes in the fetal brain at E12.5 demonstrated that these genes were enriched in terms including response to cytokine, response to decreased oxygen levels in the category of biological process. At E13.5, activating transcription factor 4 (Atf4), which is an effector of integrated stress response, was significantly upregulated in the fetal brain. Our results show that poly(I:C)-induced MIA at E12.5 leads to dysregulated neurogenesis and upregulates Atf4 in the fetal brain. These findings provide a new insight in the mechanism of MIA causing improper brain development and subsequent neurodevelopmental disorders.

Modulation of the carbohydrate-binding specificity of two Xenopus proto-type galectins by site-directed mutagenesis.

The galectin family is a representative soluble lectin group, which is responsible for the modulation of various cell functions. Although the carbohydrate-binding specificity of galectins has been well-studied, the relationship between protein structure and specificity remains to be elucidated. We previously reported the characteristics of a Xenopus laevis skin galectin, xgalectin-Va, which had diverged from galectin-1. The carbohydrate selectivity of xgalectin-Va was different from that of human galectin-1 and xgalectin-Ib (a Xenopus laevis galectin-1 homolog). In this study, we clarified the key residues for this selectivity by site-directed mutagenesis. Substitution of two amino acids of xgalectin-Va, Val56Gly/Lys76Arg, greatly enhanced the binding ability to N-acetyllactosamine and conferred significant T-cell growth inhibition activity, although the wild type had no activity. These two residues, Gly54 and Arg74 in galectin-1, would cooperatively contribute to the N-acetyllactosamine recognition. The loop region between the S4 and S5 ß-strands was involved in the binding to the TF-antigen disaccharide. The loop substitution successfully changed the carbohydrate selectivity of xgalectin-Va and xgalectin-Ib.

Decidual cells are the initial target of polyriboinosinic-polyribocytidylic acid in a mouse model of maternal viral infection.

BACKGROUND: Maternal immune activation has been implicated in the pathophysiology of neurodevelopmental disorders such as autism spectrum disorders caused by maternal infection. It has been suggested that the placental origin of inflammatory cytokines leads to neurodevelopmental disorders. However, the identity of the initial immune-activated site in the placenta, in response to maternal viral infection, is not clear. METHODS: By cross-breeding male enhanced green fluorescent protein (EGFP) transgenic mice with wild-type females, the placental tissues of maternal origin can be distinguished from those of paternal origin by EGFP expression. Using this method, at embryonic day (E) 12.5, dams were administered an intraperitoneal polyriboinosinic-polyribocytidylic acid (poly [I:C]) injection. We quantitatively analyzed the levels of phosphorylated interferon (IFN) regulatory factor 3 (pIRF3) in the placenta, and investigated the distribution of pIRF3 positive cells. RESULTS: We show that maternally derived decidual cells are the initial target of maternal poly (I:C) through the toll-like receptor 3/TIR-domain-containing the adapter-inducing interferon-ß signaling pathway. We also show that the expression of interferon-ß was upregulated in the placenta after maternal injection with poly (I:C). CONCLUSION: These results suggest that maternally derived decidual cells are the initial target of maternal poly (I:C) and that this innate immune response is likely associated with a state of maternal immune activation.

Exposure to Maternal Immune Activation Causes Congenital Unfolded Protein Response Defects and Increases the Susceptibility to Postnatal Inflammatory Stimulation in Offspring.

BACKGROUND: A number of childhood diseases have been identified, such as severe infection or autoinflammatory disease, in which immune overreaction against inflammation is a possible underlying mechanism. Previous reports have demonstrated that fetal cells exposed to maternal immune activation (MIA) induced by polyriboinosinic-polyribocytidylic acid [poly(I:C)] exhibited hypersensitivity to inflammation in vitro. However, the details of this mechanism remain unclear. Therefore, this study aimed to reveal the reaction to inflammation in offspring exposed to MIA in the prenatal period, as well as its molecular mechanism, using a viral infection mouse model. MATERIALS AND METHODS: Pregnant mice at 12.5, 14.5, and 16.5 days post coitum were injected intraperitoneally with poly(I:C) 20 mg/kg body weight (BW) or saline. Offspring aged 3-4 weeks received the second injection of 20 mg/kg BW or 4 mg/kg BW poly(I:C) or saline. Serum and tissues were collected at 2, 24, 48, and 72 h after the postnatal injection. The cytokine profile, histopathology of organs, and unfolded protein response (UPR) in offspring were examined. RESULTS: The serum levels of interleukin (IL)-6, IL-17, and interferon-γ were significantly higher in the MIA group, and acute liver necrosis was detected. Moreover, failure in UPR was observed in the MIA group compared with that in the control group. CONCLUSION: Overall, MIA exposure in utero caused failure in UPR as well as immune overreaction to the second attack of inflammation in offspring. Our results suggested that prenatal exposure to MIA might contribute to the congenital inflammatory constitution after birth.

Metabolic remodeling in the right ventricle of rats with severe pulmonary arterial hypertension.

It is generally considered that there is an increase in glycolysis in the hypertrophied right ventricle (RV) during pulmonary hypertension (PH), which leads to a decrease in glucose oxidation through the tricarboxylic acid (TCA) cycle. Although recent studies have demonstrated that fatty acid (FA) and glucose accumulated in the RV of patients with PH, the details of this remain to be elucidated. The purpose of the current study was to assess the metabolic remodeling in the RV of rats with PH using a metabolic analysis. Male rats were treated with the vascular endothelial growth factor receptor blocker SU5416 followed by 3 weeks of hypoxic conditions and 5 weeks of normoxic conditions (Su/Hx rats). Hemodynamic measurements were conducted, and the RV was harvested for the measurement of metabolites. A metabolomics analysis revealed a decreasing trend in the levels of alanine, argininosuccinic acid and downstream TCA cycle intermediates, including fumaric and malic acid and an increasing trend in branched­chain amino acids (BCAAs) in Su/Hx rats compared with the controls; however, no trends in glycolysis were indicated. The FA metabolomics analysis also revealed a decreasing trend in the levels of long­chain acylcarnitines, which transport FA from the cytosol to the mitochondria and are essential for beta­oxidation. The current study demonstrated that the TCA cycle was less activated because of a decreasing trend in the expression of fumaric acid and malic acid, which might be attributable to the expression of adenylosuccinic acid and argininosuccinic acid. These results suggest that dysregulated BCAA metabolism and a decrease in FA oxidation might contribute to the reduction of the TCA cycle reactions.

The Isoquinoline-Sulfonamide Compound H-1337 Attenuates SU5416/Hypoxia-Induced Pulmonary Arterial Hypertension in Rats.

Pulmonary arterial hypertension (PAH) is characterized by elevated pulmonary arterial pressure and right heart failure. Selective pulmonary vasodilators have improved the prognosis of PAH; however, they are not able to reverse pulmonary vascular remodeling. Therefore, a search for new treatment agents is required. H-1337 is an isoquinoline-sulfonamide compound that inhibits multiple serine/threonine kinases, including Rho-associated protein kinase (ROCK) and mammalian target of rapamycin (mTOR). Here, we investigated the effects of H-1337 on pulmonary hypertension and remodeling in the pulmonary vasculature and right ventricle in experimental PAH induced by SU5416 and hypoxia exposure. H-1337 and H-1337M1 exerted inhibitory effects on ROCK and Akt. H-1337 inhibited the phosphorylation of myosin light chain and mTOR and suppressed the proliferation of smooth muscle cells in vitro. H-1337 treatment also suppressed the phosphorylation of myosin light chain and mTOR in the pulmonary vasculature and decreased right ventricular systolic pressure and the extent of occlusive pulmonary vascular lesions. Furthermore, H-1337 suppressed aggravation of right ventricle hypertrophy. In conclusion, our data demonstrated that inhibition of ROCK and mTOR pathways with H-1337 suppressed the progression of pulmonary vascular remodeling, pulmonary hypertension, and right ventricular remodeling.

The extent of enlarged bronchial arteries is not correlated with the development of reperfusion pulmonary edema after pulmonary endarterectomy in patients with chronic thromboembolic pulmonary hypertension.

This study investigated whether dilated bronchial arteries are associated with reperfusion pulmonary edema in patients with chronic thromboembolic pulmonary hypertension. Results showed that the extent of enlarged bronchial arteries was not associated with the development of reperfusion pulmonary edema, whereas the residual pulmonary hypertension had a significant association.

Gut microbiota modification suppresses the development of pulmonary arterial hypertension in an SU5416/hypoxia rat model.

The pathogenesis of pulmonary arterial hypertension is closely associated with dysregulated inflammation. Recently, abnormal alterations in gut microbiome composition and function were reported in a pulmonary arterial hypertension experimental animal model. However, it remains unclear whether these alterations are a result or the cause of pulmonary arterial hypertension. The purpose of this study was to investigate whether alterations in the gut microbiome affected the hemodynamics in SU5416/hypoxia rats. We used the SU5416/hypoxia rat model in our study. SU5416/hypoxia rats were treated with a single SU5416 injection (30 mg/kg) and a three-week hypoxia exposure (10% O2). Three SU5416/hypoxia rats were treated with a combination of four antibiotics (SU5416/hypoxia + ABx group) for four weeks. Another group was exposed to hypoxia (10% O2) without the SU5416 treatment, and control rats received no treatment. Fecal samples were collected from each animal, and the gut microbiota composition was analyzed by 16S rRNA sequencing. The antibiotic treatment significantly suppressed the vascular remodeling, right ventricular hypertrophy, and increase in the right ventricular systolic pressure in SU5416/hypoxia rats. 16S rRNA sequencing analysis revealed gut microbiota modification in SU5416/hypoxia + ABx group. The Firmicutes-to-Bacteroidetes ratio in SU5416/hypoxia rats was significantly higher than that in control and hypoxia rats. Compared with the control microbiota, 14 bacterial genera, including Bacteroides and Akkermansia, increased, whereas seven bacteria, including Rothia and Prevotellaceae, decreased in abundance in SU5416/hypoxia rats. Antibiotic-induced modification of the gut microbiota suppresses the development of pulmonary arterial hypertension. Dysbiosis may play a causal role in the development and progression of pulmonary arterial hypertension.

The supression of DOCK family members by their specific inhibitors induces the cell fusion of human trophoblastic cells.

PURPOSE: Among the members of the DOCK family, DOCK1-5 function as guanine-nucleotide exchange factors for small GTPase Rac1, which regulates the actin cytoskeleton. It has been reported that in model organisms the Dock-Rac axis is required for myoblast fusion. We examined the role of DOCK1-5 in trophoblast fusion herein. METHODS: We used a quantitative polymerase chain reaction (qPCR) to examine the mRNA expressions of DOCK1-5 and differentiation-related genes, i.e., fusogenic genes, in human trophoblastic cell lines, BeWo and JEG-3. We treated BeWo cells with TBOPP and C21 to inhibit DOCK1 and DOCK5. Cell dynamics and cell fusion were assessed by live imaging and immunostaining. The signaling pathways induced by DOCK1/5 inhibition were examined by western blotting. RESULTS: DOCK1 and DOCK5 were expressed in BeWo cells. The inhibition of DOCK1 or DOCK5 did not prevent the cell fusion induced by forskolin (a common reagent for cell fusion); it induced cell fusion. DOCK1 inhibition induced cell death, as did forskolin. DOCK1 and DOCK5 inhibition for 24 and 48 h increased the expression of the genes ASCT2 and SYNCYTIN2, which code responsive proteins of trophoblast cell fusion, respectively. CONCLUSION: DOCK1 and DOCK5 inhibition participates in BeWo cell fusion, probably via pathways independent from forskolin-mediated pathways.

Characteristics of Japanese elderly patients with pulmonary arterial hypertension.

Previous nationwide Japanese data suggested that pulmonary arterial hypertension (PAH) predominantly affects young women. However, the number of elderly patients diagnosed with PAH has been increasing in western countries. There have been no reports on elderly PAH patients in Asian countries. This study aimed to investigate the clinical characteristics of elderly PAH patients in a Japanese cohort. Idiopathic/heritable PAH (I/H-PAH) was included in the national research project on intractable diseases. The patients were required to submit a clinical research form completed by their attending physicians. We analyzed the characteristics of Japanese I/H-PAH using the newly registered forms in 2013 (Study 1, n = 148). Also, we did a retrospective, observational cohort study at Chiba University Hospital (Study 2, n = 42). We compared the characteristics of elderly PAH patients (≥65 years old) with younger patients (<65) in both studies. Study 1 revealed a predominance of males (51% male), better hemodynamics and poorer exercise capacity in the elderly group (n = 72), compared with the younger group (n = 76) in study 1. In Study 2, elderly patients showed a male predominance (63% male), a higher ratio of smokers, a lower % carbon monoxide diffusing capacity, and poorer exercise tolerance. Elderly patients in Study 2 showed less improvement in hemodynamics with therapy. There was no significant difference in disease-specific survival between elderly and younger patients. Japanese elderly patients with I/H-PAH showed poorer exercise capacity and impaired gas exchange, but better pulmonary hemodynamics than younger patients.

Leukemia inhibitory factor induces corticotropin-releasing hormone in mouse trophoblast stem cells.

Previous studies have shown that some inflammatory cytokines promote the expression of corticotropin-releasing hormone (CRH) in trophoblasts during pregnancy and that placental CRH could induce the production of adrenocorticotropic hormone (ACTH) in humans. However, whether the same is true in rodent placenta remains unclear. In this study, we examined the effect of pro-inflammatory cytokine LIF on the induction of CRH in mouse trophoblast stem cells (mTSCs). During differentiation, the CRH levels in mTSCs gradually increased. On days 3 and 5 after LIF supplementation, Crh expression in the differentiated mTSCs was significantly increased with LIF treatment than those without LIF treatment. Moreover, the CRH concentration in the culture media increased. Thereafter, we examined the contribution of the downstream pathways of LIF to CRH induction in differentiated mTSCs. The LIF-induced upregulation of CRH was attenuated by inhibition of PI3K/AKT and MAPK phosphorylation but not by inhibition of JAK/STAT3. Therefore, in mTSCs, LIF increased Crh expression through activation of the PI3K/AKT and MAPK pathways but not by the JAK/STAT3 pathway. The present study suggests that mTSC is an ideal in vitro model for studying regulation and function of placental CRH.

Characterization of pulmonary intimal sarcoma cells isolated from a surgical specimen: In vitro and in vivo study.

Pulmonary intimal sarcoma (PIS) constitutes a rare sarcoma originating from the intimal cells of pulmonary arteries. The pathogenesis of PIS remains to be elucidated and specific treatments have not been established; therefore, prognosis is generally poor. The purpose of our study was to isolate and characterize PIS cells from a specimen resected from a patient with PIS. The surgical specimen was minced and incubated, and spindle-shaped and small cells were successfully isolated and designated as PIS-1. PIS-1 cells at passages 8-9 were used for all in vitro and in vivo experiments. Immunocytochemistry showed that PIS-1 cells were positive for vimentin, murine double minute 2, and CD44 and negative for α-smooth muscle actin, CD31, von Willebrand factor, and desmin. PIS-1 cells exhibited the hallmarks of malignant cells including the potential for autonomous proliferation, anchorage-independent growth, invasion, genetic instability, and tumorigenicity in severe combined immunodeficiency mice. The PIS-1 cells highly expressed tyrosine kinase receptors such as platelet-derived growth factor receptor, and vascular endothelial growth factor receptor 2. Pazopanib, a multi-targeted tyrosine kinase inhibitor, suppressed the proliferation of PIS-1 cells in vitro and the growth of tumors formed from xenografted PIS-1 cells. A PIS cell line was thus successfully established. The PIS-1 cells highly expressed tyrosine kinase receptors, which may be a target for treatment of PIS.

Pulmonary Hypertension that Developed During Treatment for Hepatopulmonary Syndrome and Pulmonary Arteriovenous Malformation.

Hepatopulmonary syndrome (HPS) and pulmonary arteriovenous malformation (PAVM) are hypoxemic diseases caused by right-to-left shunting but are rarely concomitant with pulmonary hypertension (PH). A 66-year-old woman with chronic hepatitis C was scheduled to undergo liver transplantation. She was referred to our department for hypoxia and an abnormal shadow in the right lung found on a preoperative examination. She was diagnosed with HPS and a PAVM in the right middle lobe. After liver transplantation, PH temporarily developed, but the pulmonary arterial pressure normalized after coil embolization. Combined HPS and PAVM may cause unique changes in pulmonary hemodynamics during treatment.