Therapeutic efficacy of intravenous infusion of mesenchymal stem cells in rat perinatal brain injury.

BACKGROUND: Perinatal brain injury is multifactorial and primarily associated with brain prematurity, inflammation, and hypoxia-ischemia. Although recent advances in perinatal medicine　have improved the survival rates of preterm infants, neurodevelopmental disorders remain a significant complication. We tested whether the intravenous infusion of mesenchymal stem cells (MSCs) had therapeutic efficacy against perinatal brain injury in rats. METHODS: Pregnant rats at embryonic day (E) 18 received lipopolysaccharide and the pups were born at E21. On postnatal day (PND) 7, the left common carotid artery of each pup was ligated, and they were exposed to 8% oxygen for 2 h. They were randomized on PND10, and MSCs or vehicle were intravenously infused. We performed behavioral assessments, measured brain volume using MRI, and performed histological analyses on PND49. RESULTS: Infused MSCs showed functional improvements in our model. In vivo MRI revealed that MSC infusion increased non-ischemic brain volume compared to the vehicle group. Histological analyses showed that cortical thickness, the number of NeuN+ and GAD67+ cells, and synaptophysin density in the non-ischemic hemisphere in the MSC group were greater than the vehicle group, but less than the control group. CONCLUSIONS: Infused MSCs improve sensorimotor and cognitive functions in perinatal brain injury and enhance neuronal growth. IMPACT: Intravenous infusion of MSCs improved neurological function in rats with perinatal brain injury, including motor, sensorimotor, cognitive, spatial, and learning memory. Infused MSCs increased residual (non-ischemic) tissue volume, number of neuronal cells, GABAergic cells, and cortical synapses in the contralesional (right) hemisphere. Intravenous administration of MSC might be suitable for the treatment of perinatal brain injury.

Intestinal Permeability of Drugs in Caco-2 Cells Cultured in Microfluidic Devices.

Microfluidic devices are attracting attention for their ability to provide a biomimetic microenvironment wherein cells are arranged in a particular pattern and provided fluidic and mechanical forces. In this study, we evaluated drug transport across Caco-2 cell layers in microfluidic devices and investigated the effects of fluid flow on drug transport and metabolism. We designed a microfluidic device that comprises two blocks of polydimethylsiloxane and a sandwiched polyethylene terephthalate membrane with pores 3.0 µm in diameter. When cultured in a dynamic fluid environment, Caco-2 cells were multilayered and developed microvilli on the surface as compared with a static environment. Drugs with higher lipophilicity exhibited higher permeability across the Caco-2 layers, as well as in the conventional method using Transwells, and the fluidic conditions had little effect on permeability. In the Caco-2 cell layers cultured in Transwells and microfluidic devices, the basal-to-apical transport of rhodamine 123, a substrate of P-glycoprotein, was greater than the apical-to-basal transport, and the presence of tariquidar, an inhibitor of P-glycoprotein, completely diminished asymmetric transport. Furthermore, fluidic conditions promoted the metabolism of temocapril by carboxylesterases. On the other hand, we showed that fluidic conditions have little effect on gene expression of several transporters and metabolic enzymes. These results provide useful information regarding the application of microfluidic devices in drug transport and metabolism studies.

Pharmacological Difference Between Platelet Aggregations in Cardioembolic Stroke Patients with Direct Oral Anticoagulants: A Pilot Study.

Background Selecting the appropriate direct oral anticoagulants (DOACs) for embolic ischemic stroke patients, especially on concurrent antiplatelet therapy, is important. However, a limited number of studies have reported on the pharmacological differences in platelet aggregation of each DOAC. We aimed to evaluate the antiplatelet effects of selected DOACs, by comparing dabigatran (a direct oral thrombin inhibitor) and factor Xa (FXa) inhibitors (apixaban and rivaroxaban) in patients who had suffered a cardioembolic stroke. Methods We retrospectively evaluated 12 patients diagnosed with a cardioembolic stroke who took any DOAC without an antiplatelet drug and underwent platelet aggregation tests within 60 days from the onset of symptoms. The platelet aggregation tests were analyzed by both light transmission aggregometry and VerifyNow®. Results Six patients (50%) took dabigatran, while the other six (50%) took an FXa inhibitor (n = 4 for apixaban and n = 2 for rivaroxaban). From the light transmission aggregometry analysis, it was found that the maximal extent of aggregation for adenosine diphosphate (ADP) was significantly higher with dabigatran than with FXa inhibitors, and the ED50 value of ADP on platelet aggregation was significantly lower with dabigatran than with FXa inhibitors. Moreover, the VerifyNow® analyses revealed that P2Y12 reaction units were significantly higher with dabigatran than with FXa inhibitors. Conclusions Dabigatran had little impact on platelet aggregation compared to FXa inhibitors in patients who had suffered a cardioembolic stroke with atrial fibrillation, and who took DOACs for secondary prevention within 60 days from the onset.

Possible role of intravenous administration of mesenchymal stem cells to alleviate interstitial cystitis/bladder pain syndrome in a Toll-like receptor-7 agonist-induced experimental animal model in rat.

BACKGROUND: Interstitial cystitis/bladder pain syndrome (IC/BPS) categorized with and without Hunner lesions is a condition that displays chronic pelvic pain related to the bladder with no efficacious treatment options. There are strong associations suggested between Hunner-type IC and autoimmune diseases. Recently, we established an animal model of Hunner-type IC using a Toll-like receptor-7 (TLR7) agonist. Intravenous infusion of mesenchymal stem cells (MSCs) can be used to treat injury via multimodal and orchestrated therapeutic mechanisms including anti-inflammatory effects. Here, we investigated whether infused MSCs elicit therapeutic efficacy associated with the TLR7-related anti-inflammatory pathway in our Hunner-type IC model. METHODS: Voiding behaviors were monitored 24 h prior to the Loxoribine (LX), which is a TLR7 agonist instillation in order to establish a Hunner-type IC model (from - 24 to 0 h) in female Sprague-Dawley rats. LX was instilled transurethrally into the bladder. At 0 h, the initial freezing behavior test confirmed that no freezing behavior was observed in any of the animals. The LX-instilled animals were randomized. Randomized LX-instilled rats were intravenously infused with MSCs or with vehicle through the right external jugular vein. Sampling tissue for green fluorescent protein (GFP)-positive MSCs were carried out at 48 h. Second voiding behavior tests were monitored from 72 to 96 h. After the final evaluation of the freezing behavior test at 96 h after LX instillation (72 h after MSC or vehicle infusion), histological evaluation with H&E staining and quantitative real-time polymerase chain reaction (RT-PCR) to analyze the mRNA expression levels of inflammatory cytokines were performed. RESULTS: Freezing behavior was reduced in the MSC group, and voiding behavior in the MSC group did not deteriorate. Hematoxylin-eosin staining showed that mucosal edema, leukocyte infiltration, and hemorrhage were suppressed in the MSC group. The relative expression of interferon-ß mRNA in the bladder of the MSC group was inhibited. Numerous GFP-positive MSCs were distributed mainly in the submucosal and mucosal layers of the inflammatory bladder wall. CONCLUSION: Intravenous infusion of MSCs may have therapeutic efficacy in a LX-instilled Hunner-type IC rat model via a TLR7-related anti-inflammatory pathway.

Clinical and Bacteriologic Analysis of Nontypeable Haemophilus influenzae Strains Isolated from Children with Invasive Diseases in Japan from 2008 to 2015.

Haemophilus influenzae type b (Hib) conjugate vaccines have led to dramatic reductions in Hib disease among young children worldwide. Nontypeable H. influenzae (NTHi) is now the major cause of invasive H. influenzae infections. We investigated the clinical characteristics of invasive NTHi diseases among children in Japan, to clarify the pathogenicity of isolated NTHi strains. The mortality rate was 10.7%, with deaths occurring mainly among children with underlying comorbidities. Biotypes II and III were the most common, and most strains (64.3%) had multiple amino acid substitutions at the Asp-350, Ser-357, Ser-385, and/or Met-377 sites of penicillin-binding protein 3. Two strains were ß-lactamase positive and ampicillin-clavulanate resistant. Biofilm indices varied widely, and IS1016 was detected in 10.7% of the strains tested. Moreover, there was wide variation in the characteristics of invasive NTHi strains. NTHi strains, showing great genetic diversity, are responsible for most invasive H. influenzae infections in children in the postvaccine era. Continuous monitoring of NTHi strains responsible for invasive diseases in children is important to detect changes in the epidemiology of invasive H. influenzae infections in the postvaccine era.

Biological relevance of tissue factor and IL-6 in arteriovenous malformations.

Arteriovenous malformations (AVMs) are congenital abnormal vessels that shunt blood directly from the arterial to the venous system without a capillary bed. The underlying pathology of AVMs is not fully understood. The objective of the study was to determine the association between the expression patterns of tissue factor (TF) and interleukin-6 (IL-6) in AVMs with clinical and pathological findings. Eighteen cases of sporadic AVM with operative specimens were included in this study. The expression of messenger RNA (mRNA) of TF and IL-6 was assayed, and association with clinical factors was investigated. The distribution of TF and IL-6 was examined with immunofluorescence. The mRNA expression of TF was significantly higher in AVM specimens than in control tissues (P = 0.002) and significantly higher in the symptomatic group than in the asymptomatic group (P = 0.037). The mRNA expression of IL-6 was likewise significantly higher in AVM specimens than in control tissues (P = 0.038). Examination of immunostained sections indicated that TF+ cells were also positive for IL-6 and were distributed around normal endothelial cells and pericytes. Moreover, TF+/IL-6+ cells also expressed CD31, vascular endothelial growth factor receptor 2 (VEGFR2), and platelet-derived growth factor receptor beta (PDGFR-beta). These results suggest that TF is elevated in AVMs and that it mediates symptomatic events. IL-6 is associated with the angiogenic activity of TF, and both are present in the same abnormal endothelial cells and pericytes. These factors may have interactive effects and may serve in a prognostic role for AVMs.

The roles of C-terminal residues on the thermal stability and local heme environment of cytochrome c' from the thermophilic purple sulfur bacterium Thermochromatium tepidum.

A soluble cytochrome (Cyt) c' from thermophilic purple sulfur photosynthetic bacterium Thermochromatium (Tch.) tepidum exhibits marked thermal tolerance compared with that from the closely related mesophilic counterpart Allochromatium vinosum. Here, we focused on the difference in the C-terminal region of the two Cyts c' and examined the effects of D131 and R129 mutations on the thermal stability and local heme environment of Cyt c' by differential scanning calorimetry (DSC) and resonance Raman (RR) spectroscopy. In the oxidized forms, D131K and D131G mutants exhibited denaturing temperatures significantly lower than that of the recombinant control Cyt c'. In contrast, R129K and R129A mutants denatured at nearly identical temperatures with the control Cyt c', indicating that the C-terminal D131 is an important residue maintaining the enhanced thermal stability of Tch. tepidum Cyt c'. The control Cyt c' and all of the mutants increased their thermal stability upon the reduction. Interestingly, D131K exhibited narrow DSC curves and unusual thermodynamic parameters in both redox states. The RR spectra of the control Cyt c' exhibited characteristic bands at 1,635 and 1,625 cm(-1), ascribed to intermediate spin (IS) and high spin (HS) states, respectively. The IS/HS distribution was differently affected by the D131 and R129 mutations and pH changes. Furthermore, R129 mutants suggested the lowering of their redox potentials. These results strongly indicate that the D131 and R129 residues play significant roles in maintaining the thermal stability and modulating the local heme environment of Tch. tepidum Cyt c'.

Intravenous Preload of Mesenchymal Stem Cells Rescues Erectile Function in a Rat Model of Cavernous Nerve Injury.

INTRODUCTION: We evaluated the potential preventive effects and mechanisms of intravenously preloaded mesenchymal stem cells (MSCs) for erectile dysfunction (ED) in a cavernous nerve (CN) injury model. METHODS: Male Sprague-Dawley (SD) rats were used for this study. Rats were randomized into two groups. One group was intravenously preloaded with MSCs (1.0 × 10(6) cells in 1 mL total fluid volume) and the other was infused with medium alone (1 mL Dulbecco's modified Eagle's medium [DMEM]) for sham control, respectively. Crushed CN injury was induced immediately after infusion. The surgeon was blind to the experimental conditions (MSC or medium). MAIN OUTCOME MEASURES: To assess erectile function, we measured the intracavernous pressure (ICP) and arterial pressure (AP) at 1 hour and 2 weeks after CN injury. After measuring the initial ICP/AP of pre-injury (normal) male SD rats, they were randomized into the two groups and infused with MSCs or medium. PKH26-labelled MSCs were used for tracking. To investigate the mRNA expression levels of neurotrophins in the major pelvic ganglia (MPG), we performed real-time quantitative real-time polymerase chain reaction. RESULTS: The reduction of ICP/AP and area under the curve of ICP (ICP-AUC) in the MSC group was significantly lower than in the DMEM group (P < 0.05; P < 0.05) at 1 hour. The ICP/AP and ICP-AUC at 2 weeks post-injury in the MSC group was significantly higher than in the DMEM group (P < 0.01; P < 0.05). The preloaded PKH26-labelled MSCs were detected in the MPG and CN using confocal microscopy indicating homing of the cells to the injured nerve and ganglia. Glia cell-derived neurotrophic factor (GDNF) and neurturin, which are important neurotrophic factors for erection, had expression levels in MPG significantly higher in the MSC group than in the DMEM group (P < 0.01, 0.05). CONCLUSION: Intravenous preload of MSCs before a CN injury may prevent or reduce experimental ED.

Age-related changes in the diurnal variation of ketogenesis in patients with type 2 diabetes and relevance to hypoglycemic medications.

To assess the significance of ketogenesis in the management of diabetes mellitus, we analyzed the factors associated with the diurnal variation of the plasma ketone body levels. The subjects consisted of 220 patients with type 2 diabetes, aged 60 ± 15 years, without advanced complications. They ate a standardized, low-fat meal at 8:00, 12:00, and 18:00. The plasma levels of 3-hydroxybutyrate (3HB) and free fatty acid (FFA) were increased before breakfast and before dinner. The plasma glucose concentration was almost the same at any blood sampling time point among age quartiles. However, the 3HB levels were significantly decreased with age, which was most obvious before dinner. The FFA levels also decreased with age, but the decline was mild. A multiple regression analysis with stepwise selection revealed that age was an independent, negative contributor and that the pre-breakfast FFA concentration was an independent, positive contributor to the pre-breakfast 3HB levels. Regarding the pre-dinner 3HB levels, in addition to age and the pre-dinner FFA concentration, the uses of sulfonylurea and dipeptidyl peptidase-4 inhibitors were independent negative contributors. The metabolism of ketone bodies is an alternative energy source for the brain under conditions of starvation. While excessive ketogenesis leads to critical ketoacidosis, inadequate ketone body production could be associated with a propensity to develop neurohypoglycemia in elderly patients treated with insulin secretagogues. Because age-related changes in ketogenesis were the most significant before dinner, attention should be paid not only to fasting but also to the pre-dinner levels of 3HB.

Perfluoropolyether-Based Gut-Liver-on-a-Chip for the Evaluation of First-Pass Metabolism and Oral Bioavailability of Drugs.

In the evolving field of drug discovery and development, multiorgans-on-a-chip and microphysiological systems are gaining popularity owing to their ability to emulate in vivo biological environments. Among the various gut-liver-on-a-chip systems for studying oral drug absorption, the chip developed in this study stands out with two distinct features: incorporation of perfluoropolyether (PFPE) to effectively mitigate drug sorption and a unique enterohepatic single-passage system, which simplifies the analysis of first-pass metabolism and oral bioavailability. By introducing a bolus drug injection into the liver compartment, hepatic extraction alone could be evaluated, further enhancing our estimation of intestinal availability. In a study on midazolam (MDZ), PFPE-based chips showed more than 20-times the appearance of intact MDZ in the liver compartment effluent compared to PDMS-based counterparts. Notably, saturation of hepatic metabolism at higher concentrations was confirmed by observations when the dose was reduced from 200 µM to 10 µM. This result was further emphasized when the metabolism was significantly inhibited by the coadministration of ketoconazole. Our chip, which is designed to minimize the dead volume between the gut and liver compartments, is adept at sensitively observing the saturation of metabolism and the effect of inhibitors. Using genome-edited CYP3A4/UGT1A1-expressing Caco-2 cells, the estimates for intestinal and hepatic availabilities were 0.96 and 0.82, respectively; these values are higher than the known human in vivo values. Although the metabolic activity in each compartment can be further improved, this gut-liver-on-a-chip can not only be used to evaluate oral bioavailability but also to carry out individual assessment of both intestinal and hepatic availability.

Multiple intravenous infusions versus a single infusion of mesenchymal stem cells in a rat model of cerebral ischemia.

OBJECTIVE: Recent randomized clinical trials of a single infusion of mesenchymal stem cells (MSCs) for acute cerebral stroke revealed a limited functional recovery outcome. Conversely, animal studies suggest that multiple MSC infusions may enhance functional recovery by inducing neural plasticity, which indicates that a multiple-infusion approach might be effective for stroke treatment in humans. The objective of this study was to investigate whether multiple infusions of MSCs enhance functional outcomes during the acute phase of cerebral ischemia. METHODS: Rats subjected to permanent middle cerebral artery occlusion (MCAO) were randomized into four groups: 1) vehicle group (infusion of vehicle only), 2) MSC-1 group (single administration of the standard MSC dose on day 3), 3) high-dose MSC group (single administration of three times the standard MSC dose on day 3), and 4) MSC-3 group (multiple administrations of the standard MSC dose on days 3, 10, and 17). MSCs were administered via the femoral vein. Behavioral performance and ischemic lesion volume were examined using in vivo MRI every 7 days from day 3 to day 45 after MCAO induction. The thickness of the corpus callosum (CC) was determined using Nissl staining, and the area of the CC was measured using ex vivo MRI. Interhemispheric connections within the CC were assessed using ex vivo MRI diffusion tensor imaging. RESULTS: The MSC-3 group exhibited the most significant motor recovery and increased CC thickness and area among all groups. Increased CC thickness and area were correlated with improved behavioral function 45 days after MCAO induction. Neural tracts through interhemispheric connections via the CC were most pronounced in the MSC-3 group, and this anatomical change showed a positive relationship with behavioral function. CONCLUSIONS: Multiple infusions of MSCs led to histological changes in the CC and neural tracts within the CC. These results indicate that multiple systemic infusions of MSCs had a greater beneficial effect in the acute phase of MCAO than a single standard or high-dose infusion of MSCs.

Rehabilitation facilitates functional improvement following intravenous infusion of mesenchymal stem cells in the chronic phase of cerebral ischemia in rats.

The primary objective of this study was to investigate the potential facilitating effects of daily rehabilitation for chronic cerebral ischemia following the intravenous infusion of mesenchymal stem cells (MSC) in rats. The middle cerebral artery (MCA) was occluded by intraluminal occlusion using a microfilament (MCAO). Eight weeks after MCAO induction, the rats were used as a chronic cerebral ischemia model. Four experimental groups were studied: Vehicle group (medium only, no cells); Rehab group (vehicle + rehabilitation), MSC group (MSC only); and Combined group (MSC + rehabilitation). Rat MSCs were intravenously infused eight weeks after MCAO induction, and the rats received daily rehabilitation through treadmill exercise for 20 min. Behavioral testing, lesion volume assessment using magnetic resonance imaging (MRI), and histological analysis were performed during the observation period until 16 weeks after MCAO induction. All treated animals showed functional improvement compared with the Vehicle group; however, the therapeutic efficacy was greatest in the Combined group. The combination therapy is associated with enhanced neural plasticity shown with histological analysis and MRI diffusion tensor imaging. These findings provide behavioral evidence for enhanced recovery by combined therapy with rehabilitation and intravenous infusion of MSCs, and may form the basis for the development of clinical protocols in the future.

Intravenous Infusion of Autologous Mesenchymal Stem Cells Expanded in Auto Serum for Chronic Spinal Cord Injury Patients: A Case Series.

Objective: The safety, feasibility, and potential functional improvement following the intravenous infusion of mesenchymal stem cells (MSCs) were investigated in patients with chronic severe spinal cord injury (SCI). Methods: The intravenous infusion of autologous MSCs cultured in auto-serum under Good Manufacturing Practices (GMP) was administered to seven patients with chronic SCI (ranging from 1.3 years to 27 years after the onset of SCI). In addition to evaluating feasibility and safety, neurological function was evaluated using the American Spinal Injury Association Impairment Scale (AIS), International Standards for Neurological Classification of Spinal Cord Injury (ISCSCI-92), and Spinal Cord Independence Measure III (SCIM-III). Results: No serious adverse events occurred. Neither CNS tumors, abnormal cell growth, nor neurological deterioration occurred in any patients. While this initial case series was not blinded, significant functional improvements and increased quality of life (QOL) were observed at 90 and 180 days post-MSC infusion compared to pre-infusion status. One patient who had an AIS grade C improved to grade D within six months after MSC infusion. Conclusions: This case series suggests that the intravenous infusion of autologous MSCs is a safe and feasible therapeutic approach for chronic SCI patients. Furthermore, our data showed significant functional improvements and better QOL after MSC infusion in patients with chronic SCI. A blind large-scale study will be necessary to fully evaluate this possibility.

Prevalence and clinical characteristics of unremembered nocturnal eating in diabetic subjects: Kurume sleep trouble in obesity and metabolic disorders (KUSTOMED) study.

Nighttime food intake is associated with weight gain and higher HbA1c levels. We experienced night eaters who have no memory of their nocturnal eating in the morning. In this study, the curious night eating behavior was designated as "unremembered nocturnal eating syndrome (UNES)". We screened 1,169 patients with diabetes for sleep quality and abnormal eating behavior at night using the Pittsburgh Sleep Quality Index questionnaire with an additional question regarding UNES. When abnormal nocturnal eating behavior was noted, detailed clinical information was extracted from interviews with the patients. We identified 9 patients who experienced UNES. They had a higher BMI compared with subjects who reported no such episodes. Among them, 6 patients who consumed food at night without memory 2-5 times per month or more had significantly higher HbA1c levels. Continuous glucose monitoring in a patient with type 1 diabetes revealed an abrupt elevation of glucose levels from midnight when some foods were consumed. Eight of the 9 patients were taking benzodiazepine and/or non-benzodiazepine hypnotic agents when they experienced the episodes. The prevalence of UNES was 0.8% in all subjects and 4% in those taking hypnotic drugs. The ratio of hypnotic drug use in subjects with UNES was significantly higher than for individuals without UNES (89% vs. 17%, p<0.0001). Although UNES seems to be etiologically heterogeneous, hypnotics-induced parasomnia and/or anterograde amnesia may be associated with the behavior. UNES is not rare in diabetic patients on hypnotic medicine and may be a hidden cause of unexpected morning hyperglycemia.

A conditional gene expression system in Porphyromonas gingivalis for study of the secretion mechanisms of lipoproteins and T9SS cargo proteins.

The Gram-negative anaerobe, Porphyromonas gingivalis, is known to be a pathogen associated with chronic periodontitis. P. gingivalis possesses virulence factors such as fimbriae and gingipain proteinases. Fimbrial proteins are secreted to the cell surface as lipoproteins. In contrast, gingipain proteinases are secreted into the bacterial cell surface via the type IX secretion system (T9SS). The transport mechanisms of lipoproteins and T9SS cargo proteins are entirely different and remain unknown. Therefore, using the Tet-on system developed for the genus Bacteroides, we newly created a conditional gene expression system in P. gingivalis. We succeeded in establishing conditional expression of nanoluciferase and its derivatives for lipoprotein export, of FimA for a representative of lipoprotein export, and of T9SS cargo proteins such as Hbp35 and PorA for representatives of type 9 protein export. Using this system, we showed that the lipoprotein export signal, which has recently been found in other species in the phylum Bacteroidota, is also functional in FimA, and that a proton motive force inhibitor can affect type 9 protein export. Collectively, our conditional protein expression method is useful for screening inhibitors of virulence factors, and may be used to investigate the role of proteins essential to bacterial survival in vivo.

Actin Alpha 2, Smooth Muscle (ACTA2) Is Involved in the Migratory Potential of Malignant Gliomas, and Its Increased Expression at Recurrence Is a Significant Adverse Prognostic Factor.

Malignant glioma is a highly invasive tumor, and elucidating the glioma invasion mechanism is essential for developing novel therapies. We aimed to highlight actin alpha 2, smooth muscle (ACTA2) as potential biomarkers of brain invasion and distant recurrence in malignant gliomas. Using the human malignant glioma cell line, U251MG, we generated ACTA2 knockdown (KD) cells treated with small interfering RNA, and the cell motility and proliferation of the ACTA2 KD group were analyzed. Furthermore, tumor samples from 12 glioma patients who underwent reoperation at the time of tumor recurrence were utilized to measure ACTA2 expression in the tumors before and after recurrence. Thereafter, we examined how ACTA2 expression correlates with the time to tumor recurrence and the mode of recurrence. The results showed that the ACTA2 KD group demonstrated a decline in the mean motion distance and proliferative capacity compared to the control group. In the clinical glioma samples, ACTA2 expression was remarkably increased in recurrent samples compared to the primary samples from the same patients, and the higher the change in ACTCA2 expression from the start to relapse, the shorter the progression-free survival. In conclusion, ACTA2 may be involved in distant recurrence in clinical gliomas.

A practical protocol for high-spatial-resolution magnetic resonance angiography for cerebral arteries in rats.

BACKGROUND: Magnetic resonance angiography (MRA) is an important tool in rat models of cerebrovascular disease. Although MRA has long been used in rodents, the image quality is typically not as high as that observed in clinical practice. Moreover, studies on MRA image quality in rats are limited. This study aimed to develop a practical high-spatial-resolution MRA protocol for imaging cerebral arteries in rats. NEW METHOD: We used the "half position method" regarding coil placement and modified the imaging parameters and image reconstruction method. We applied this new imaging method to measure maturation-related signal changes on rat MRAs. RESULTS: The new practical high-spatial-resolution MRA imaging protocol obtained a signal intensity up to 3.5 times that obtained using a basic coil system, simply by modifying the coil placement method. This method allowed the detection of a gradual decrease in the signal in cerebral vessels with maturation. COMPARISON WITH EXISTING METHODS: A high-spatial-resolution MRA for rats was obtained with an imaging time of approximately 100 min. Comparable resolution and image quality were obtained using the new protocol with an imaging time of 30 min CONCLUSIONS: The new practical high-spatial-resolution MRA protocol can be implemented simply and successfully to achieve high image quality with an imaging time of approximately 30 min. This protocol will benefit researchers performing MRA imaging in cerebral artery studies in rats.

Complete genome sequence of Mycoplasma pneumoniae type 2a strain 309, isolated in Japan.

Mycoplasma pneumoniae strain 309, a type 2a (subtype 2 variant) strain of this bacterium, has variations in the P1 protein, which is responsible for attachment of the bacterium to host cells. Here, we report the complete genome sequence of M. pneumoniae strain 309 isolated from a pneumonia patient in Japan.

Behavioral analysis of Drosophila transformants expressing human taste receptor genes in the gustatory receptor neurons.

Transgenic Drosophila expressing human T2R4 and T2R38 bitter-taste receptors or PKD2L1 sour-taste receptor in the fly gustatory receptor neurons and other tissues were prepared using conventional Gal4/UAS binary system. Molecular analysis showed that the transgene mRNAs are expressed according to the tissue specificity of the Gal4 drivers. Transformants expressing the transgene taste receptors in the fly taste neurons were then studied by a behavioral assay to analyze whether transgene chemoreceptors are functional and coupled to the cell response. Since wild-type flies show strong aversion against the T2R ligands as in mammals, the authors analyzed the transformants where the transgenes are expressed in the fly sugar receptor neurons so that they promote feeding ligand-dependently if they are functional and activate the neurons. Although the feeding preference varied considerably among different strains and individuals, statistical analysis using large numbers of transformants indicated that transformants expressing T2R4 showed a small but significant increase in the preference for denatonium and quinine, the T2R4 ligands, as compared to the control flies, whereas transformants expressing T2R38 did not. Similarly, transformants expressing T2R38 and PKD2L1 also showed a similar preference increase for T2R38-specific ligand phenylthiocarbamide (PTC) and a sour-taste ligand, citric acid, respectively. Taken together, the transformants expressing mammalian taste receptors showed a small but significant increase in the feeding preference that is taste receptor and also ligand dependent. Although future improvements are required to attain performance comparable to the endogenous robust response, Drosophila taste neurons may serve as a potential in vivo heterologous expression system for analyzing chemoreceptor function.

Long-term efficacy of sitagliptin for the treatment of type 2 diabetic patients in Japan.

We studied the efficacy of sitagliptin in type 2 diabetic patients of our outpatient clinics. Since December in 2009, 164 patients have been treated by sitagliptin for their management of diabetes. HbA1c decreased by 0.8% in all patients without any change in mean body weight after 3 months. However, actually HbA1c did not decrease in 30 patients, and more than half of patients showed weight gain to some extent. Patients were classified according to the reduction of HbA1c and analyzed based on this category. Baseline characters such as age, gender, duration of diabetes, BMI, concomitantly used oral hypoglycemic agents and the score for life-style assessment were not related to glucose-lowering effect of sitagliptin. Ninety eight patients whose HbA1c had decreased after 3 months were further followed-up for another 3 months. Among them 45 patients showed some relapsing of HbA1c after 6 months, and they were compared with 53 patients without relapsing. More cases had been switched from α-glucosidase inhibitor (α-GI) and the score for life-style assessment was lower in relapsing patients compared to those in patients without relapsing. In conclusion, clinicians should keep the fact in mind that the individual variation of glucose-lowering effects and the possibility of relapsing exist during sitagliptin treatment, and that concern about life-style is still a quite important issue to prevent weight gain and the relapsing of blood glucose control.