Involvement of TRPV1-ANO1 Interactions in Pain-Enhancing Mechanisms.

Primary sensory neurons detect potentially dangerous environmental situations via many "sensor" proteins located on the plasma membrane. Although receptor-type cation channels are thought to be the major sensors in sensory neurons, anion channels are also important players in the peripheral nervous system. Recently, we showed that transient receptor potential vanilloid 1 (TRPV1) interacts with anoctamin 1 (ANO1, also called TMEM16A) in primary sensory neurons and that this interaction enhanced TRPV1-mediated pain sensation. In that study, we induced ANO1 currents by application of capsaicin to small DRG neurons and showed that ANO1-dependent depolarization following TRPV1 activation could evoke more action potentials. Furthermore, capsaicin-evoked pain-related behaviors in mice were strongly inhibited by a selective ANO1 blocker. Together these findings indicate that selective ANO1 inhibition can reduce pain sensation. We also investigated non-specific inhibitory effects on ion channel activities to control ion dynamics via the TRPV1-ANO1 complex. We found that 4-isopropylcyclohexanol (4-iPr-CyH-OH) had an analgesic effect on burning pain sensations through its inhibition of TRPV1 and ANO1 together. Additionally, 4-iPr-CyH-OH did not have clear agonistic effects on TRPV1, TRPA1, and ANO1 activity individually. These results indicate that 4-iPr-CyH-OH could function globally to mediate TRP-ANO1 complex functions to reduce skin hypersensitivity and could form the basis for novel analgesic agents.

The relationship between stroke severity and prior direct oral anticoagulant therapy in patients with acute ischaemic stroke and non-valvular atrial fibrillation.

BACKGROUND AND PURPOSE: Anticoagulant treatment with a vitamin K antagonist (VKA) has been reported to reduce stroke severity when patients with atrial fibrillation (AF) suffer acute ischaemic stroke (AIS). Direct oral anticoagulant (DOAC) therapy also has the potential to reduce the initial severity of AIS. However, the effect of DOAC therapy on the severity of AIS is not well known. The aim of the present study was to investigate the effect of DOACs on initial stroke severity in patients with AIS and non-valvular AF. METHODS: From March 2011 to July 2016, consecutive patients with AIS having non-valvular AF were recruited. The effects of prior DOAC treatment on severity were assessed by multivariate logistic regression analyses. RESULTS: A total of 484 patients [208 women; median age 79 (interquartile range, 71-85) years; National Institutes of Health Stroke Scale (NIHSS) score 9 (interquartile range, 3-20)] were enrolled. Of these, 352 (73%) were on no anticoagulant medication, 54 (11%) were undertreated with a VKA, 35 (7%) were sufficiently treated (admission prothrombin time-international normalized ratio: ≥2.0 for patients <70 years old and ≥1.6 for ≥70 years old) with a VKA and 43 (9%) were on a DOAC. The initial NIHSS score (median 10 in patients with no anticoagulation, 13 in undertreated VKA, 7 in sufficient VKA and 6 in DOAC, P = 0.018) was different among the groups. Multivariate analysis showed that DOAC was independently and negatively associated with severe (initial NIHSS score ≥ 10) stroke (odds ratio, 0.39; P = 0.041), compared with no anticoagulant therapy. CONCLUSIONS: Direct oral anticoagulant treatment prior to the event should reduce initial stroke severity in patients with AIS and non-valvular AF.

Predictors of therapeutic effect of transarterial chemoembolisation using drug-eluting beads for hepatocellular carcinoma.

AIMS: To identify predictors of a therapeutic effect after transarterial chemoembolisation using drug-eluting beads (DEB-TACE) for hepatocellular carcinoma (HCC). MATERIALS AND METHODS: Between January 2015 and July 2015, tumour variables and angiographic data were collected for 25 patients (49 target lesions) after they had undergone the DEB-TACE procedure for HCC. The therapeutic effect was evaluated according to the Response Evaluation Criteria in Cancer of the Liver at follow-up dynamic computed tomography (CT) performed within 1-4 months after the procedure. A p<0.05 was considered significant. RESULTS: On a target lesion basis, the objective response (TE3/4) rate was 63.3% (31 of 49). On univariate analysis, larger size (≥2 cm) was a predictor of an objective response (p=0.029). The tumour location of the medial (segment 4) or caudate (segment 1) lobe also indicated a poor therapeutic effect (TE1/2), but not at the level of significance (p=0.051). Multivariate analysis identified tumour size (odds ratio, 8.60; 95% confidence interval, 1.87-62.8) and tumour location (odds ratio, 12.2; 95% confidence interval, 2.12-129.8) as significant factors associated with a therapeutic effect. On a patient basis, 10 of 25 (40%) patients showed complete response/partial response. There were no significant differences between complete response/partial response and stable disease/progressive disease regarding age, gender, tumour markers, history of previous treatment, Child-Pugh class, T-stage, or Barcelona Clinic Liver Cancer Staging. CONCLUSION: A short-term therapeutic effect was associated with tumour size and location on a target lesion basis.

Heterogeneity of non-cancerous liver parenchyma on gadoxetic acid-enhanced MRI: an imaging biomarker for hepatocellular carcinoma development in chronic liver disease.

AIM: To clarify whether the heterogeneity of non-cancerous liver parenchyma (NLP) in the hepatobiliary phase on gadoxetic acid enhanced magnetic resonance imaging (MRI) is correlated with hepatocellular carcinoma (HCC) development. MATERIALS AND METHODS: Institutional review board approval was obtained, and the requirements for informed consent were waived for this retrospective study. The imaging characteristics of 84 patients with chronic liver disease who underwent gadoxetic acid-enhanced 3T MRI between January 2013 and October 2014 were examined retrospectively. For the evaluation of the heterogeneity of the intensity in the hepatobiliary phase, the largest possible region of interest was placed on the NLP, and the skewness and kurtosis were calculated using ImageJ software. Skewness is the degree of asymmetry of a histogram, and kurtosis is a measure of the peak. Based on the median values of kurtosis and skewness, the patients were classified into four categories and the categories were compared between the 49 patients with HCC (HCC group) and the 35 patients without HCC (non-HCC group). RESULTS: Kurtosis was significantly higher in the HCC group compared to the non-HCC group (1.19±1.15 versus 0.43±0.83; p=0.0006). Skewness was significantly lower in the HCC group than in the non-HCC group (1.19±1.15 versus 0.43±0.83; p=0.0152). In a multivariate logistic analysis, the category showing lower-than-the-median (-0.1185) skewness and higher-than-the-median (0.547) kurtosis was significantly and independently associated with HCC development (p=0.0031). CONCLUSION: The heterogeneity of NLP in the hepatobiliary phase on gadoxetic acid enhanced MRI may reflect the development of HCC.

Fatty change in moderately and poorly differentiated hepatocellular carcinoma on MRI: a possible mechanism related to decreased arterial flow.

AIM: To clarify the frequency of fatty change in moderately and poorly differentiated hepatocellular carcinomas (mHCCs and pHCCs) and its relationship to arterial blood flow. MATERIALS AND METHODS: One hundred and thirty-six surgically resected HCC lesions were studied. All patients had undergone dynamic magnetic resonance imaging (MRI) with chemical-shift-encoded water-fat imaging (CSI). The presence of fat was identified by a signal drop-off on CSI and confirmed at pathology. Lesions were classified into four groups in the arterial phase; G1, hypointense; G2, isointense; G3, slightly and heterogeneously hyperintense; G4, markedly and homogeneously hyperintense. The number of cumulative arteries (CAs) in the tumours in the pathology examination were counted. RESULTS: A fat component was observed significantly more frequently in the pHCCs (13/21; 61.9%) compared to the mHCCs (32/101; 31.7%; p=0.013). The numbers of lesions in each group were as follows: (G1, G2, G3, G4) = (18, 9, 23, 4) in the HCCs with fat; (1, 6, 24, 51) in the HCCs without fat (p<0.001); (5, 5, 18, 4) in the mHCCs with fat; (0, 3, 19, 47) in the mHCCs without fat (p<0.001); (11, 0, 2, 0) in the pHCCs with fat; (0, 2, 3, 3) in the pHCCs without fat (p=0.001). The number of CAs in the fat-containing HCCs (5.5±2.9) was significantly lower than that in the HCCs without fat (10.8±5.3; p<0.001). CONCLUSION: A fat component was more commonly observed in the pHCCs than in the mHCCs. The present results showed a possible mechanism of fatty change in mHCCs and pHCCs in relation to decreased arterial blood supply.

Balanced MR cholangiopancreatography with motion-sensitised driven-equilibrium (MSDE) preparation: feasibility of Gd-EOB-DTPA-enhanced biliary examination.

AIM: To evaluate the effectiveness of motion-sensitised driven-equilibrium (MSDE)-prepared balanced magnetic resonance cholangiopancreatography (MRCP) in a gadolinium ethoxybenzyl diethylene triamine pentaacetic acid (Gd-EOB-DTPA)-enhanced study compared to conventional T2-weighted MRCP. MATERIALS AND METHODS: Fifteen patients (seven male and eight female patients) prospectively underwent conventional three-dimensional turbo spin-echo T2-weighted MRCP and MSDE-balanced MRCP using a 1.5 T MRI system after hepatobiliary phase image acquisition. For quantitative evaluation, the contrast-to-noise ratio (CNR) of the common hepatic duct to liver tissue was calculated. For qualitative analysis, two radiologists evaluated the depiction of the biliary system and main pancreatic duct (MPD) using a scoring system. Signal suppression of the portal vein (PV) and hepatic vein (HV) on MSDE-balanced MRCP was also scored. RESULTS: MSDE-balanced MRCP showed significantly higher CNR than T2-weighted MRCP. For all biliary structures, the mean depiction scores of MSDE-balanced MRCP were significantly higher than those of T2-weighted MRCP, whereas the mean depiction score of MPD with MSDE-balanced MRCP was significantly lower than that of T2-weighted MRCP. Signal suppression of the PV and HV was thought to be clinically sufficient. CONCLUSIONS: MSDE-balanced MRCP more clearly depicted biliary structures compared with T2-weighted MRCP in a Gd-EOB-DTPA-enhanced study. This sequence may be utilised for routine MRCP on Gd-EOB-DTPA-enhanced MRI.

Detectability of hepatocellular carcinoma on gadoxetic acid-enhanced MRI at 3 T in patients with severe liver dysfunction: clinical impact of dual-source parallel radiofrequency excitation.

AIM: To clarify the detectability of hepatocellular carcinoma (HCC) on gadoxetic acid-enhanced MRI at 3 T with dual-source parallel radiofrequency (RF) excitation. MATERIALS AND METHODS: Twelve patients with 26 HCCs who each underwent multidetector row CT (MDCT), gadoxetic acid-enhanced MRI with dual-source parallel RF excitation, and angiography-assisted CT prior to living related-liver transplantation. Three blinded readers independently reviewed the images obtained by each imaging technique for the presence of HCC on a segment-by-segment basis using a five-point confidence scale. The area under the receiver operating characteristic curve (Az), sensitivity, and specificity were compared among the three techniques. RESULTS: The Az values of gadoxetic acid-enhanced MRI were highest for all readers, although no significant difference in Az value among the three methods was obtained. No significant differences in sensitivity or specificity were observed among the three techniques for each reader. CONCLUSION: Gadoxetic acid-enhanced MRI at 3 T with dual-source parallel RF excitation has relatively high-level diagnostic potential for the detection of HCC in patients with severe liver dysfunction, which was equivalent to that of MDCT and angiography-assisted CT. Dual-source parallel RF excitation would have a clinical impact on 3 T MRI of the liver.

MR Imaging Detection of CNS Lesions in Tuberous Sclerosis Complex: The Usefulness of T1WI with Chemical Shift Selective Images.

BACKGROUND AND PURPOSE: CNS lesions of tuberous sclerosis complex are diagnosed mainly by T2WI, FLAIR, and sometimes T1WI with magnetization transfer contrast. The usefulness of T1WI with chemical shift selective images was recently reported in focal cortical dysplasia type IIb, which has histopathologic and imaging features similar to those of tuberous sclerosis complex. We investigated the usefulness of the T1WI with chemical shift selective images in detecting CNS lesions of tuberous sclerosis complex. MATERIALS AND METHODS: We retrospectively reviewed 25 consecutive patients with tuberous sclerosis complex (mean age, 11.9 [SD, 8.9] years; 14 males) who underwent MR imaging including T1WI, T1WI with magnetization transfer contrast, T1WI with chemical shift selective, T2WI, and FLAIR images. Two neuroradiologists assessed the number of CNS lesions in each sequence and compared them in 2 steps: among T1WI, T1WI with magnetization transfer contrast and T1WI with chemical shift selective images, and among T2WI, FLAIR, and T1WI with chemical shift selective images. We calculated the contrast ratio of the cortical tubers and of adjacent normal-appearing gray matter and the contrast ratio of radial migration lines and adjacent normal-appearing white matter in each sequence and compared them. RESULTS: T1WI with chemical shift selective images was significantly superior to T1WI with magnetization transfer contrast for the detection of radial migration lines and contrast ratio of radial migration lines. There was no significant difference between T1WI with chemical shift selective images and T1WI with magnetization transfer contrast for the detection of cortical tubers and the contrast ratio of the cortical tubers. Both T2WI and FLAIR were statistically superior to T1WI with chemical shift selective images for the detection of cortical tubers. T1WI with chemical shift selective images was significantly superior to T2WI and FLAIR for the detection of radial migration lines. CONCLUSIONS: The usefulness of T1WI with chemical shift selective images in detecting radial migration lines was demonstrated. Our findings suggest that the combination of T1WI with chemical shift selective images, T2WI, and FLAIR would be useful to evaluate the CNS lesions of patients with tuberous sclerosis complex in daily clinical practice.

A foodborne outbreak of a group A streptococcal infection in a Japanese university hospital.

We describe an outbreak of foodborne tonsillopharyngitis caused by group A streptococcus (GAS), a rarely reported event that occurred during a campus orientation meeting in Japan. Of 461 students and staff members who had eaten boxed lunches during a meeting at Kitasato University, 298 developed sore throat and/or fever, and 285 underwent medical examination. Amoxicillin was prescribed when throat culture specimens yielded GAS. The attack rate was 64.6%. T-25 GAS was isolated from 150 examined persons. Of 65 patients who received amoxicillin for 3 days, GAS was eradicated before the first follow-up throat culture in 46 (70.8%) cases. Susceptibility was demonstrated to penicillins, cephalosporins, and macrolides in 86 GAS isolates obtained more than once from a given patient. GAS strains isolated at various time points were indistinguishable by pulsed-field gel electrophoresis (PFGE), and prtF1 was present. GAS strains were often difficult to eradicate because of a short initial treatment period, patient compliance problems, and the presence of prtF1.

Effects of raloxifene and hormone replacement therapy on forearm skin elasticity in postmenopausal women.

OBJECTIVES: Hormone replacement therapy (HRT) increases skin elasticity in postmenopausal women. However, the effects of raloxifene, a selective estrogen receptor modulator (SERM), on skin degenerative changes in postmenopausal women remain unknown. We investigated whether raloxifene increases skin elasticity, similar to HRT, in postmenopausal women. METHODS: In a 12-month trial, 17 postmenopausal women (mean age, 66.4+/-7.8 years) received continuous raloxifene treatment (60 mg/day), 19 women (56.2+/-6.4 years) received continuous 17-beta estradiol treatment using a patch (0.72 mg/2 days) plus cyclic medroxyprogesterone acetate (2.5mg/day, for 12 days/month), and 11 women (58.1+/-7.3 years) did not receive either therapy. In each subject, the skin elasticity of the forearm was measured using a suction device at baseline and at 12 months after the start of the study. RESULTS: Raloxifene and HRT significantly increased skin elasticity from 52.4+/-3.8% and 64.1+/-7.2% at baseline to 55.1+/-4.7% and 67.4+/-7.4% after 12 months, respectively (P<0.05, each), but the untreated subjects did not exhibit any significant change in skin elasticity during the study. The delta value for skin elasticity was significantly higher among the raloxifene and HRT subjects than among the untreated subjects (P<0.05, each). CONCLUSIONS: These findings suggest that raloxifene may have a beneficial effect on skin elasticity, which undergoes degenerative changes in postmenopausal women, in addition to its effects on bone metabolism.

Radiologic and Pathologic Features of the Transmantle Sign in Focal Cortical Dysplasia: The T1 Signal Is Useful for Differentiating Subtypes.

BACKGROUND AND PURPOSE: The transmantle sign is a characteristic MR imaging finding often seen in focal cortical dysplasia type IIb. The transmantle sign is typically hyperintense on T2WI and FLAIR and hypointense on T1WI. However, in some cases, it shows T1 high signal. We evaluated the imaging and pathologic findings to identify the causes of the T1 high signal in the transmantle sign. MATERIALS AND METHODS: We retrospectively reviewed the preoperative imaging data of 141 consecutive patients with histologically proved focal cortical dysplasia. We selected 25 patients with focal cortical dysplasia with the transmantle sign and divided them into groups based on the pathologic focal cortical dysplasia subtype and T1 signal of the transmantle sign. We evaluated the clinical, radiologic, and pathologic findings, including the number of balloon cells and dysmorphic neurons and the severity of gliosis or calcifications and compared them among the groups. RESULTS: Nine of the 25 patients had a T1-high-signal transmantle sign; the other 16 patients did not. All 9 patients with a T1-high-signal transmantle sign were diagnosed as type IIb (group A). Of the 16 patients with no T1-high-signal transmantle sign, 13 were diagnosed as having type IIb (group B), and the other 3 patients, as type IIa (group C). The number of balloon cells was significantly higher in group A than in the other groups, but there were no differences regarding dysmorphic neurons, the severity of gliosis, or calcifications. CONCLUSIONS: Approximately 6% (9/141) of this patient series had a T1-high-signal transmantle sign, and all were type IIb. The signal may reflect a rich density of balloon cells. This finding could support the differentiation of subtypes, especially type IIb.

Clinicopathological investigation of odontogenic fibroma in tuberous sclerosis complex.

Tuberous sclerosis complex (TSC) is an autosomal dominant inherited disease characterized by systemic hamartoma and diverse systemic features. TSC1 and TSC2 are the causative genes, and mental retardation, epileptic seizures, and facial angiofibroma develop in many patients with the disease. The case of a patient with TSC who developed a central odontogenic fibroma of the mandible is reported here. The patient was a 21-year-old woman who was referred with a swelling of the labial gingiva in the region of the right lower lateral incisor and canine. Dental radiography revealed a multilocular radiolucent region with a clear boundary. The right lower lateral incisor and canine were continuous with the lesion and thus were excised en bloc. The lesion was encapsulated and easily dissected. The diagnosis on immunohistological staining was odontogenic fibroma without an epithelial component. TSC1/2 gene mutation causes abnormal activation of mammalian target of rapamycin (mTOR) downstream of the PI3K-AKT pathway. The odontogenic fibroma in this patient was positive for mTOR, suggesting that the development of the odontogenic fibroma was the result of abnormal activation of mTOR, as in angiofibroma. The clinical course of this patient is presented and the developmental mechanism of central odontogenic fibroma is discussed.

Does Neuroticism Influence Oral Health-Related QOL in Patients with Removable Partial Dentures?

This prospective cohort study investigated the relationship between patient neuroticism and oral health-related quality of life (OHRQoL) before and after prosthetic treatment as well as changes in OHRQoL-namely, treatment efficacy. Sixty-three patients (23 men and 40 women; mean age 67.2 ± 8.6 years), who were scheduled to receive new removable partial dentures (RPDs), were recruited. OHRQoL was assessed using the Japanese version of the Oral Health Impact Profile (OHIP-J). The Japanese version of the NEO Five-Factor Inventory (NEO-FFI) was used to assess neuroticism. Spearman's rank correlation coefficient was calculated to determine the association between neuroticism and OHIP-J scores before and after treatment. After stratifying patients according to neuroticism score, the Wilcoxon signed-rank test was used for intragroup comparison of OHIP-J scores before and after treatment. Moreover, logistic regression analysis was used to determine the impact of covariates on treatment efficacy such as age, sex, Eichner classification, neuroticism, changes in maximal occlusal force, and OHIP-J scores before treatment. Statistical analyses showed that higher neuroticism scores were associated with higher total OHIP-J scores before treatment ( r = 0.41, P = 0.001) but were not associated with OHIP-J scores after treatment ( r = 0.07, P = 0.566). When the effect of all independent variables was analyzed in multivariate analysis, neuroticism and OHIP-J scores before treatment affected treatment efficacy. These results suggest that OHRQoL of patients with higher levels of neuroticism was low before prosthetic treatment but significantly improved by oral rehabilitation with RPDs to the same level as patients with lower levels of neuroticism. Knowledge Transfer Statement: The results of this study may change the clinical perception of the effect of prosthetic rehabilitation with removable partial dentures in patients with higher levels of neuroticism. The study concluded that prosthetic rehabilitation could contribute toward satisfaction even in neurotic patients, who are presumed to show less satisfaction with their oral status.

Prevalence of, and risk factors for, carriage of carbapenem-resistant Enterobacteriaceae among hospitalized patients in Japan.

BACKGROUND: The prevalence of carbapenem-resistant Enterobacteriaceae (CRE) has been reported to be lower in Japan than in many other countries. However, extensive surveillance for CRE carriage has not been performed in Japan. AIM: To investigate the prevalence of CRE carriage in Japan among convalescent patients considered to be at high risk of being CRE carriers using an improved selective culture medium. METHODS: A cross-sectional survey was conducted in 22 acute care hospitals (ACHs) and 21 long-term care hospitals (LTCHs) in northern Osaka from December 2015 to January 2016. Patients who used incontinence aids, an enteral feeding tube or a urinary catheter were enrolled. Faecal specimens were examined using the newly developed M-ECC for imipenemase (IMP)-producing CRE, which is the most prevalent form of CRE in Japan. The positive isolates were analysed by polymerase chain reaction and sequencing. Risk factors associated with carriage were analysed by logistic regression. FINDINGS: Among 1507 patients, 184 (12.2%) carried CRE. The percentage of positive patients was significantly higher in LTCHs (14.9%) than in ACHs (3.6%) (P<0.001). Risk factors for CRE carriage were longer hospital stay [odds ratio (OR) 2.59; 95% confidence interval (CI) 1.87-3.60], enteral feeding (OR 3.03, 95% CI 2.08-4.42) and antibiotic exposure (OR 2.00, 95% CI 1.40-2.87). Among the 233 CRE isolates identified, 223 were IMP producers; the remaining isolates did not produce carbapenemase. CONCLUSIONS: This is the first Japanese report to demonstrate the significant spread of CRE in both ACHs and LTCHs using an improved selective medium. A coordinated regional approach may help to prevent further spread.

Elevated arterial stiffness in postmenopausal women with osteoporosis.

OBJECTIVES: Osteoporosis and increased pulse wave velocity (PWV) are cardiovascular risk factors. We investigated the relationship between PWV and bone mass in the lumbar spine in postmenopausal women. METHODS: We studied the PWV in 95 women; 38 postmenopausal women with normal spinal bone mineral density (BMD), 32 osteopenic postmenopausal women, and 25 osteoporotic postmenopausal women. The brachial-ankle PWV (baPWV) was measured using an automated device. The BMD of the lumbar spine (L2-L4) was measured using dual-energy X-ray absorptiometry. RESULTS: After adjusting for age and years since menopause, women with osteoporosis had a significantly higher baPWV than those with normal BMD (1500 +/- 220 cm/s versus 1340 +/- 215 cm/s; P < 0.05), but no significant differences in baPWV were seen between the osteoporotic and osteopenic groups or between the osteopenic and normal BMD groups. In univariate regression analysis, the baPWV was significantly negatively correlated with BMD (r = -0.450, P < 0.01), and significantly positively correlated with age (r = 0.601, P < 0.01), years since menopause (r = 0.577, P < 0.01), systolic blood pressure (r = 0.295, P < 0.01), and diastolic blood pressure (r = 0.264, P < 0.05), but was not with other variables. In multivariate regression analysis, the baPWV was significantly correlated with BMD (P < 0.05), but not with other variables. CONCLUSIONS: Postmenopausal women with osteoporosis may have elevated arterial stiffness, suggesting that osteoporotic postmenopausal women may have a higher risk of cardiovascular disease.

Insulin-like growth factor I rapidly induces tyrosine phosphorylation of a Mr 150,000 and a Mr 160,000 protein in highly metastatic mouse colon carcinoma 26 NL-17 cells.

Insulin-like growth factor I (IGF-I) stimulates the proliferation of highly metastatic NL-17 cells to a greater extent than poorly metastatic NL-44 cells, both of which are derived from mouse colon carcinoma 26. The NL-17 cells have been compared with NL-44 cells for the signal transduction pathway of IGF-I. IGF-I receptors of both cell types were identified by affinity labeling, and there was no significant difference between the two cell types in the amount or the autophosphorylation activity of the IGF-I receptors. However, when IGF-I-dependent tyrosine phosphorylation of cellular components was examined, remarkable tyrosine phosphorylation of proteins with molecular weights of 150,000 (pp150) and 160,000 (pp160) was found in NL-17 cells. In contrast, this phosphorylation stayed at significantly lower levels in NL-44 cells than in NL-17 cells. The phosphorylation of pp150 and pp160 was induced within 10 s after the addition of IGF-I and reached its maximal level by 30 s. After the removal of IGF-I, the phosphorylation of pp150 and pp160 was reduced to the basal level within 30 min. This phosphorylation was not induced by platelet-derived or epidermal growth factor. The pp150 and pp160 were not absorbed by wheat germ agglutinin-agarose. They were found in the soluble fraction of cytoplasm but not in the membrane or the cytoskeleton. The pp150 and pp160 might be endogenous substrates of IGF-I receptor kinase. These results suggest that tyrosine phosphorylation of pp150 and pp160 mediates the higher proliferative response of NL-17 cells to IGF-I.

Lactobacillus rhamnosus GG increases Toll-like receptor 3 gene expression in murine small intestine ex vivo and in vivo.

Administration of Lactobacillus rhamnosus GG (LGG) has been reported to be therapeutically effective against acute secretory diarrhoea resulting from the structural and functional intestinal mucosal lesions induced by rotavirus infection; however, the underlying mechanisms remain to be completely elucidated. Because Toll-like receptor 3 (TLR3) plays a key role in the innate immune responses following the recognition of rotavirus, the present study examined whether LGG influences TLR3 gene expression in murine small intestine ex vivo and in vivo. We employed cultured intestinal organoids derived from small intestinal crypts as an ex vivo tissue model. LGG supplementation increased TLR3 mRNA levels in the intestinal organoids, as estimated by quantitative real-time polymerase chain reaction. Likewise, single and 7-day consecutive daily administrations of LGG increased TLR3 mRNA levels in the small intestine of C57BL/6N mice. The mRNA levels of other TLRs were not substantially altered both ex vivo and in vivo. In addition, LGG supplementation increased the mRNA levels of an antiviral type 1 interferon, interferon-α (IFN-α), and a neutrophil chemokine, CXCL1, upon stimulation with a synthetic TLR3 ligand, poly(I:C) in the intestinal organoids. LGG administration did not alter IFN-α and CXCL1 mRNA levels in the small intestine in vivo. Supplementation of other bacterial strains, Bifidobacterium bifidum and Lactobacillus paracasei, failed to increase TLR3 and poly(I:C)-stimulated CXCL1 mRNA levels ex vivo. We propose that upregulation of TLR3 gene expression may play a pivotal role in the therapeutic efficacy of LGG against rotavirus-associated diarrhoea. In addition, we demonstrated that intestinal organoids may be a promising ex vivo tissue model for investigating host-pathogen interactions and the antiviral action of probiotics in the intestinal epithelium.

Tumor progression following transformation of murine monocytes by v-myc: acquisition of immortalization and tumorigenicity.

Monocyte transformation by the v-myc oncogene has been used to study myelomonocytic tumor progression in vitro. Murine monocytes transformed by a recombinant retrovirus containing MC29 v-myc were found to exhibit a proliferative burst to day 28-40 post-infection. There-after growth slowed and cell number remained relatively static to day 80-90 post-infection. During both the proliferative and quiescent periods, the cells were dependent on the myelomonocytic growth factor CSF-1 for growth and viability. Analysis of this transformation revealed that the initial transformants were polyclonal, non-immortal, and non-tumorigenic in syngeneic mice. At day 80-90 post infection, a fresh round of cellular proliferation occurred and, in contrast to the initial burst, growth was sustained allowing the establishment of cell lines. These lines were found to be monoclonal, immortal, growth factor independent and, in certain cases, tumorigenic in syngeneic mice. Associated with the establishment of growth factor independent cell lines was the constitutive synthesis of the myelomonocytic growth factor, CSF-1. Proto-oncogene screening of the initial transformants and the cell lines also revealed the expression of c-raf and the CSF-1 receptor, c-fms. Our results indicate that, following transformation by v-myc, monocytes can progress in vitro to become growth factor independent and immortal and that both monocyte transformation and immortalization can be dissociated from tumorigenicity.

Transformation of early erythroid precursor cells (BFU-E) by a recombinant murine retrovirus containing v-erb-B.

Avian erythroblastosis virus (AEV) is a replication-defective retrovirus that transforms erythroid and fibroblast cells in vitro and in vivo. The transforming ability of AEV is due primarily to the oncogene v-erb-B. A recombinant murine retrovirus has been constructed by inserting a chimeric gag-v-erb-B gene into a Moloney murine leukemia virus based vector. This retrovirus was used to examine v-erb-B-induced transformation of murine hematopoietic cells. Infection of murine primary fetal liver, adult bone marrow or adult spleen cells with the recombinant virus generated large hemoglobinized erythroid colonies in the absence of exogenous growth factors. Generation of such colonies usually requires the presence of erythropoietin (Epo) and interleukin-3 (IL-3). These growth-factor independent colonies were shown to be derived from early (BFU-E) and not late (CFU-E) erythroid progenitor cells, and the effect was not attributable to growth factors elicited by the virus-producing cell lines. In order to confirm that the recombinant virus was responsible for this transformation of BFU-E to growth factor independence, bone marrow cells from post 5-fluorouracil treated mice were infected and used to repopulate lethally-irradiated mice. Growth factor-independent BFU-E were obtained in up to 30% of day-13 spleen colonies and it was shown by DNA analysis that cells from these colonies contained integrated provirus. Our results indicate that v-erb-B transforms early erythroid progenitors to growth factor independent growth and subsequent differentiation to erythrocytes -a process that normally requires Epo plus either IL-3 or granulocyte-macrophage colony stimulating factor (GM-CSF).

Activity of midbrain reticular formation and neocortex during the progression of human non-rapid eye movement sleep.

To clarify the neural correlates and brain activity during the progression of human non-rapid eye movement (NREM) sleep, we examined the absolute regional cerebral blood flow (rCBF) during light and deep NREM sleep and during wakefulness in normal humans using positron emission tomography with H(2)(15)O. Relative changes in rCBF during light and deep NREM sleep in comparison to the rCBF during wakefulness were also analyzed. During light NREM sleep, the rCBF in the midbrain, in contrast to that in the pons and thalamic nuclei, did not decrease when compared to that during wakefulness, whereas rCBF decreased in the left medial frontal gyrus, left inferior frontal gyrus, and left inferior parietal gyrus of the neocortex. During deep NREM sleep, the rCBF in the midbrain tegmentum decreased, and there was a marked and bilateral decrease in the rCBF in all neocortical regions except for the perirolandic areas and the occipital lobe. There have been three groups of brain structures, each representing one type of deactivation during the progression of NREM sleep. The activity of the midbrain reticular formation is maintained during light NREM sleep and therefore represents a key distinguishing characteristic between light and deep NREM sleep. Selective deactivation of heteromodal association cortices, including those related to language, occurs with increasingly deep NREM sleep, which supports the recent theory that sleep is not a global, but it is a local process of the brain.