Three-dimensional reversed fast imaging with steady-state precession diffusion-weighted imaging for the detection of middle ear cholesteatoma.

AIM: To determine the usefulness of three-dimensional reversed fast imaging with steady-state precession diffusion-weighted imaging (3D-PSIF DWI) for the detection of middle ear cholesteatoma. MATERIALS AND METHODS: The study population consisted of 81 patients who underwent 3D-PSIF-DWI at 3 T. They included cholesteatoma in 73 cases, otitis media in five, and cholesterol granuloma in three. Two observers independently performed qualitative evaluations for the detection of cholesteatoma and measured apparent diffusion coefficient (ADC) values and ADC ratios of the lesions. Kappa (κ) statistics, the intraclass correlation coefficient (ICC), the independent t-test, and receiver operating characteristic (ROC) analysis were used for statistical analysis. Pair-wise comparison of the ROC curves was performed using the area under the ROC curve (AUC). RESULTS: Interobserver agreement and ICC for the qualitative and quantitative evaluations were excellent (κ=0.92 and ICC=0.90-0.92, respectively). The ADC value and the ADC ratio were significantly lower for cholesteatoma than non-cholesteatoma lesions (p<0.0001). In <5 mm cholesteatoma group, the diagnostic performance of the ADC value (AUC=0.97) and the ADC ratio (AUC=1) was significantly superior to qualitative 3D-PSIF-DWI (AUC=0.76; p=0.0001 and <0.0001, respectively). For ≥5 mm cholesteatoma group, there were no significant differences in diagnostic performance among the three parameters. CONCLUSION: 3D-PSIF-DWI sequence is useful for the detection of middle ear cholesteatomas, especially <5 mm lesions.

Coping strategies and risk of cardiovascular disease incidence and mortality: the Japan Public Health Center-based prospective Study.

AIMS: Coping strategies may be significantly associated with health outcomes. This is the first study to investigate the association between baseline coping strategies and cardiovascular disease (CVD) incidence and mortality in a general population cohort. METHODS AND RESULTS: The Japan Public Health Center-based prospective Study asked questions on coping in its third follow-up survey (2000-04). Analyses on CVD incidence and mortality included 57 017 subjects aged 50-79 without a history of CVD and who provided complete answers on approach- and avoidance-oriented coping behaviours and strategies. Cox regression models, adjusted for confounders, were used to determine hazard ratios (HRs) according to coping style. Mean follow-up time was 7.9 years for incidence and 8.0 years for mortality.The premorbid use of an approach-oriented coping strategy was inversely associated with incidence of stroke (HR = 0.85; 95% CI, 0.73-1.00) and CVD mortality (HR = 0.74; 95% CI, 0.55-0.99). Stroke subtype analyses revealed an inverse association between the approach-oriented coping strategy and incidence of ischaemic stroke (HR = 0.79; 95% CI, 0.64-0.98) and a positive association between the combined coping strategy and incidence of intra-parenchymal haemorrhage (HR = 2.03; 95% CI, 1.01-4.10). Utilizing an avoidance coping strategy was associated with increased mortality from ischaemic heart disease (IHD) only in hypertensive individuals (HR = 3.46; 95% CI, 1.07-11.18). The coping behaviours fantasizing and positive reappraisal were associated with increased risk of CVD incidence (HR = 1.24; 95% CI, 1.03-1.50) and reduced risk of IHD mortality (HR = 0.63; 95% CI, 0.40-0.99), respectively. CONCLUSION: An approach-oriented coping strategy, i.e. proactively dealing with sources of stress, may be associated with significantly reduced stroke incidence and CVD mortality in a Japanese population-based cohort.

UCP1 genetic polymorphism (-3826 A/G) diminishes resting energy expenditure and thermoregulatory sympathetic nervous system activity in young females.

BACKGROUND: Recent findings regarding the existence of functional brown adipose tissue (BAT) in adult humans suggest a physiological role of BAT and uncoupling protein 1 (UCP1)-linked thermogenesis in energy balance. OBJECTIVE: To investigate whether UCP1 polymorphism was associated with resting energy expenditure (REE) and thermoregulatory sympathetic nervous system (SNS) activity in humans. METHODS: A total of 82 healthy females (20-22 years) were genotyped for the -3826 A/G polymorphism of the UCP1 gene using a fluorescent allele-specific DNA primer assay system. REE was measured by indirect calorimetry. The thermoregulatory SNS activity was assessed by heart rate variability power spectral analysis according to our previously reported method. Each subject was studied in the morning, after an overnight fast. Nutritional values were calculated on the basis of 2-day food records. RESULTS: The frequencies of A/A, A/G and G/G genotypes were 0.27, 0.45 and 0.28, respectively. No significant difference was found in anthropometric indexes among the three groups. However, in the G/G group, the percentage of energy consumed as fat was lower (A/A: 30.7 ± 1.1%, A/G: 31.3 ± 1.0%, G/G: 26.0 ± 1.2%, P<0.01), and energy intake tended to be lower (A/A: 7209 ± 310 kJ d(-1), A/G: 7075 ± 280 kJ d(-1), G/G: 6414 ± 264 kJ d(-1), P=0.16). With regard to metabolic parameters, group differences were observed in REE (A/A: 5599 ± 170 kJ d(-1), A/G: 5054 ± 115 kJ d(-1), G/G: 4919 ± 182 kJ d(-1), P<0.01) and in thermoregulatory SNS activity (A/A: 313 ± 47 ms(2), A/G: 333 ± 42 ms(2), G/G: 185 ± 23 ms(2), P<0.05). CONCLUSION: Diminished REE in G-allele carriers as well as reduced thermoregulatory SNS activity for the G/G genotype, suggest that attenuated UCP1-linked thermogenesis has an adverse effect on the regulation of energy balance.

Effects of Rice Wine Lees on Cognitive Function in Community-Dwelling Physically Active Older Adults: A Pilot Randomized Controlled Trial.

BACKGROUND: Rice wine lees (RWL), a Japanese traditional fermented product, is a rich source of one-carbon metabolism-related nutrients, which may have beneficial effects on cognitive function. OBJECTIVES: We aimed to examine the effect of the RWL on cognitive function in community-dwelling physically active older adults. DESIGN: Double-blind, randomized, placebo-controlled study (clinical trial number: UMIN 000027158). SETTING: Community-based intervention including assessments conducted at the University of Hyogo and a public liberal arts school in Himeji City, Japan. PARTICIPANTS: A total of 35 community-dwelling older adults (68-80 years) who performed mild exercise before and during the trial were assigned to either the RWL (n=17) or the placebo group (n=18). INTERVENTION: Daily consumption of 50 g RWL powder, which contained one-carbon metabolism-related nutrients, or the placebo powder (made from soy protein and dextrin) for 12 weeks. Both supplements included equivalent amounts of energy and protein. MEASUREMENTS: Montreal Cognitive Assessment, computerized cognitive function test, and measurements of serum predictive biomarkers (transthyretin, apolipoprotein A1, and complement C3) were conducted at baseline and follow-up. RESULTS: Visual selective attention and serum transthyretin significantly improved in the RWL group, whereas there was no significant change in the placebo group. No significant group difference was observed in the remaining cognitive performance tests. CONCLUSIONS: RWL supplements seem to have a few effects on cognitive function in community-dwelling physically active older adults. However, the impact was limited; therefore, further studies with sufficient sample size are warranted to elucidate this issue.

Embryonic lethality of molecular chaperone hsp47 knockout mice is associated with defects in collagen biosynthesis.

Triple helix formation of procollagen after the assembly of three alpha-chains at the C-propeptide is a prerequisite for refined structures such as fibers and meshworks. Hsp47 is an ER-resident stress inducible glycoprotein that specifically and transiently binds to newly synthesized procollagens. However, the real function of Hsp47 in collagen biosynthesis has not been elucidated in vitro or in vivo. Here, we describe the establishment of Hsp47 knockout mice that are severely deficient in the mature, propeptide-processed form of alpha1(I) collagen and fibril structures in mesenchymal tissues. The molecular form of type IV collagen was also affected, and basement membranes were discontinuously disrupted in the homozygotes. The homozygous mice did not survive beyond 11.5 days postcoitus (dpc), and displayed abnormally orientated epithelial tissues and ruptured blood vessels. When triple helix formation of type I collagen secreted from cultured cells was monitored by protease digestion, the collagens of Hsp47+/+ and Hsp47+/- cells were resistant, but those of Hsp47-/- cells were sensitive. These results indicate for the first time that type I collagen is unable to form a rigid triple-helical structure without the assistance of molecular chaperone Hsp47, and that mice require Hsp47 for normal development.

Adiponectin gene single-nucleotide polymorphisms and treatment response to obesity.

BACKGROUND: In the adiponectin gene polymorphisms, single-nucleotide polymorphism (SNP)-45 and SNP276 have reportedly been associated with obesity, Type 2 diabetes, and other features of metabolic syndrome. AIM: Whether these adiponectin SNP affect obesity-related parameters during caloric restriction in obese subjects. SUBJECTS AND METHODS: Thirty- two obese Japanese women were treated by meal replacement with a low calorie diet for 8 weeks and asked to maintain their habitual lifestyle. Obesity-related parameters were measured before and after the treatment period. We determined four SNP (T45G, I164T, G276T, and C-11377G) using a fluorescent allele-specific DNA primer assay systemand FRET probe assay system. RESULTS: After the treatment, the extent of decrease in waist circumference was greater in the subjects with the G/G or G/T genotype of SNP276 than in those with the T/T genotype (p=0.026). As for SNP45, the extent of decrease in triglyceride levels was greater in the subjects with the T/T genotype than in those with the T/G genotype (p=0.003). For SNP-11377, the extent of decrease in systolic blood pressure and fasting plasma glucose was greater in the subjects with the C/G or G/G genotype than in those with the C/C genotype (p=0.044). CONCLUSION: Our findings indicate that each SNP in the adiponectin gene might modify the change in obesity-related parameters during meal replacement with a low calorie diet.

Stromal cells promote bone invasion by suppressing bone formation in ameloblastoma.

AIMS: To study the stromal variation and role of stromal-tumour cell interaction in impaired bone formation as well as enhanced bone resorption in ameloblastoma. METHODS AND RESULTS: Four types of stroma were observed histologically; fibrous, desmoplastic, myxoid and myxoid with hyalinization. Osteoblast and osteoclast were counted using haematoxylin and eosin sections and immunohistochemistry with CD68. After histomorphometric analysis, only fibrous and myxoid types of stroma were distinctly identified. Secreted frizzled-related peptide (sFRP)-2, transforming growth factor-beta 1 and receptor activator of nuclear factor-kappaB ligand (RANKL) revealed strong expression in myxoid type compared with the normal stroma. Bone morphogenetic protein (BMP)-2 was negative in myxoid type, but positive in normal stroma. Fibrous-type stroma showed weak expression of all antigens except RANKL compared with myxoid type. CONCLUSIONS: The results suggest that stroma does not act only in bone resorption, but also in the suppression of new bone formation. sFRP-2 is the main factor for impaired bone formation. The expression of markers related to osteoclastogenesis and suppression of osteoblast formation is higher in myxoid-type than in fibrous-type stroma. Tumour cells create a favourable environment for impaired bone formation by secreting sFRP-2 as well as bone resorption by secreting RANKL and interleukin-6.

Deletion mapping of chromosome 4q22-35 and identification of four frequently deleted regions in head and neck cancers.

Head and neck squamous cell carcinoma (HNSCC) is a diverse group of cancers that are frequently aggressive in their biologic behavior. Inactivation of tumor suppressor gene (TSG) is one of the most critical steps leading to HNSCC. Loss of heterozygosity analysis is very sensitive method for the detection of frequent allelic loss in a chromosomal locus. This method has been considered as an important evidence for the localization of TSGs. We analyzed loss of heterozygosity (LOH) at chromosome 4q22-35 region by using 14 polymorphic microsatellite markers in 83 matched normal and HNSCC tissues. LOH was detected at least in one location in 71 of 83 (86%) tumor tissues. Frequent deletions were detected at the location of microsatellite markers, D4S2909 (46%), D4S2623 (51%), D4S406 (48%), D4S1644 (45%) and D4S2979 (40%). Four different frequently deleted regions at 4q22, 4q25, 4q31 and 4q34-35 were observed. These regions include several putative TSGs such as Caspase-6, SMARCAD1, SMARCA5, SAP30 and ING2. Further molecular analysis of each gene should be performed to clarify their roles in head and neck squamous cell carcinogenesis.

Notch signaling in benign and malignant ameloblastic neoplasms.

BACKGROUND: In general, Notch is a representative signal which controls morphosis and differentiation of cells, but its role in human odontogenic neoplasms, especially in ameloblastoma and its malignant counterpart, ameloblastic carcinoma, is not known. METHODS: We examined Notch1 peptide and its gene (mRNA) in an ameloblastoma (case 1: 27-year-old female, right mandibular tumor) and an ameloblastic carcinoma (case 2: 93-year-old female, right mandibular tumor), using immunohistochemistry (IHC) and in situ hybridization (ISH) techniques. RESULTS: Notch1 intracellular domain (NICD) positive products were observed in the cells at the peripheral layer of most proliferating epithelial tumor nests in case 1. In case 2, positive products were similarly detected. In particular, small numbers of mitoses were identified in the nuclear region with intense NICD positive reaction. CONCLUSIONS: Notch signaling plays some role in cytological differentiation or acquisition of tissue specific characteristics in neoplastic cells of odontogenic neoplasms, including ameloblastoma and ameloblastic carcinoma. Notch1 may also contribute to cell cycle arrest induced by Notch1 activation in ameloblastic carcinoma.

Gene expression of Jagged2 in mandibular condylar cartilage development.

Expression pattern of Jagged2 gene in mandibular condylar cartilage was examined by means of in situ hybridization (ISH) technique. At E14, Jagged2 mRNA signals appeared in cytoplasm of proliferating chondrocytes. From E15 to E19, Jagged2 mRNA was detected throughout almost all cytoplasm in all layers. However, the distribution pattern was not uniform. These results suggest that Jagged2 plays an essential role for mandibular condylar cartilage morphogenesis and development.

Lack of B-RAF mutations in head and neck squamous cell carcinoma.

B-RAF is one of the most commonly mutated oncogenes in human cancer. However, the mutation status of B-RAF has not been established completely in HNSCC. We have analysed the mutation status of the kinase domain of the B-RAF gene (exons 11 and 15) in 91 Japanese HNSCC patients as well as 12 HNSCC cell lines. DNA was extracted and amplified by PCR. Mutations were then analysed by SSCP mutation detection method. Since V600EB-RAF constitutes 90 % of the mutations identified in B-RAF in human cancers, we also used MASA analysis to specifically detect this mutation in exon 15 of B-RAF. Using both methods, no mutation was found in both exon 11 and 15 in all patients and cell lines. Mu tations are absent or rare in the kinase domain of B-RAF in Japanese HNSCC. However, more studies are still needed to determine its usefulness as a target for molecular therapy in these patients.

Effect of combined bifidobacteria supplementation and resistance training on cognitive function, body composition and bowel habits of healthy elderly subjects.

Physical exercise exerts favourable effects on brain health and quality of life of the elderly; some of these positive health effects are induced by the modulation of microbiota composition. We therefore conducted a randomised, double blind, placebo-controlled trial that assessed whether a combination of Bifidobacterium spp. supplementation and moderate resistance training improved the cognitive function and other health-related parameters in healthy elderly subjects. Over a 12-week period, 38 participants (66-78 years) underwent resistance training and were assigned to the probiotic Bifidobacterium supplementation (n=20; 1.25×1010 cfu each of Bifidobacterium longum subsp. longum BB536, B. longum subsp. infantis M-63, Bifidobacterium breve M-16V and B. breve B-3) or the placebo (n=18) group. At baseline and at 12 weeks, we assessed the cognitive function, using the Japanese version of the Montreal Cognitive Assessment instrument (MoCA-J); modified flanker task scores; depression-anxiety scores; body composition; and bowel habits. At 12 weeks, the MoCA-J scores showed a significant increase in both the groups, while the flanker task scores of the probiotic group increased more significantly than those of the placebo group (0.35±0.9 vs -0.29±1.1, P=0.056). Only the probiotic group showed a significant decrease in the depression-anxiety scores (5.2±6.3 to 3.4±5.5, P=0.012) and body mass index (24.0±2.8 to 23.5±2.8 kg/m2, P<0.001), with a significant increase in the defecation frequency (5.3±2.3 to 6.4±2.3 times/5 days, P=0.023) at 12 weeks. Thus, in healthy elderly subjects, combined probiotic bifidobacteria supplementation and moderate resistance training may improve the mental condition, body weight and bowel movement frequency.

Plasminogen activator inhibitor-1 contributes to the deleterious effect of obesity on the outcome of thrombotic ischemic stroke in mice.

BACKGROUND: It is widely accepted that obesity is a risk factor for ischemic heart disease, but the association with stroke is less clear. Adipose tissue is an important source of plasminogen activator inhibitor-1 (PAI-1), the main inhibitor of plasminogen activation. OBJECTIVE: To test the hypothesis that elevated PAI-1 levels associated with obesity negatively affect the outcome of thrombotic ischemic stroke. METHODS: Middle cerebral artery (MCA) occlusion was induced photochemically in mice with nutritionally induced or genetically determined obesity and their lean counterparts. RESULTS: The MCA occlusion time (to obtain complete occlusion) was significantly shorter in obese (nutritionally induced) than in lean wild-type (WT) C57Bl/6 mice, whereas the infarct size was significantly larger and intracranial hemorrhage (ICH) was enhanced (all P < 0.05). Similar observations were made in genetically obese ob/ob mice, as compared to lean WT littermates. In both strains, obesity was associated with markedly elevated circulating PAI-1 levels, probably originating from the fat tissue. In contrast, PAI-1-deficient lean and obese mice did not display significant differences in MCA occlusion time, infarct volume or ICH. CONCLUSIONS: Plasminogen activator inhibitor-1 may play a functional role in the deleterious effect of obesity on the outcome of thrombotic ischemic stroke in mice.

Stromelysin-1 (MMP-3) is critical for intracranial bleeding after t-PA treatment of stroke in mice.

BACKGROUND: Tissue-type plasminogen activator (t-PA) is approved for treatment of ischemic stroke patients, but it may increase the risk of intracranial bleeding (ICB). Matrix metalloproteinases (MMPs), which can be activated through the plasminogen/plasmin system, may contribute to ICB after ischemic stroke. OBJECTIVES: To explore the contribution of plasminogen, MMP-3 and MMP-9 to ICB associated with t-PA treatment after ischemic stroke. METHODS: Using a thrombotic middle cerebral artery occlusion (MCA-O) model, ICB was studied in mice with genetic deficiencies of plasminogen (Plg(-/-)), stromelysin-1 (MMP-3(-/-)), or gelatinase B (MMP-9(-/-)) and their corresponding wild-type (WT) littermates. The induction of MMP-3 and MMP-9 was also studied in C57BL/6 WT mice. RESULTS: ICB induced by t-PA (10 mg kg(-1)) was significantly less than WT in Plg(-/-) (P < 0.05) and MMP-3(-/-) (P < 0.05) but not in MMP-9(-/-) mice. Furthermore, administration of the broad-spectrum MMP inhibitor GM6001 after t-PA treatment reduced ICB significantly (P < 0.05) in MMP-3(+/+) mice, but had no effect on MMP-3(-/-) mice. MMP-3 expression was significantly enhanced at the ischemic hemisphere; with placebo treatment, it was expressed only in neurons, whereas it was up-regulated in endothelial cells with t-PA treatment. Although MMP-9 expression was also significantly enhanced at the ischemic brain, the amount and the distribution were comparable in mice with and without t-PA treatment. CONCLUSIONS: Our data with gene-deficient mice thus suggest that plasminogen and MMP-3 are relatively more important than MMP-9 for the increased ICB induced by t-PA treatment of ischemic stroke.

Notch signaling in mandibular condylar cartilage development.

OBJECTIVE: The purpose of this study was to investigate the expression pattern of Notch signaling in mandibular condylar cartilage, as a type of secondary cartilage. METHODS: Mandibular condyle of ddY mice were fixed from embryonic day 14 (E14) through just after birth (equivalent to E19). Samples were cut into 4 mum serial sections through the central area of the mandibular condyle at the sagittal plane. Serial sections were examined using histological, immunohistochemical (IHC) and in situ hybridization (ISH) techniques. RESULTS: At E14, there were no developmental features of mandibular condyle. At the distal upper portion of developmental mandibular bone, mesenchymal cell proliferation and condensation without metacholomatic reaction to toluidine blue (TB) were seen. At E15, mandibular condylar cartilage was clearly evident, as TB metacholomasia. In IHC specimens at E14, expression of Notch1 intracellular domain (NICD) was observed in the nuclei of coagulating mesenchymal cells. After E15, NICD appeared in the nuclei and the cytoplasms of cells. In ISH examination at E14, expressions of Notch1 mRNA appeared in cytoplasm of proliferating chondrocytes. From E15 to E19, Notch1 mRNA was detected throughout almost all cytoplasm in all layers. CONCLUSION: These IHC and ISH results suggest that Notch signaling plays an essential role for mandibular condylar cartilage morphogenesis and development.

In vitro cell culture model for anti-cataract drug penetration studies.

Immortalized human corneal epithelial cells (HCECs) and human lens epithelial cells (HLECs) were cultured in vitro. Cells were observed under a phase-contrast microscope and the integrity of cell monolayers was assayed by transepithelial electrical resistance (TEER) determination. The permeability of disulfiram (DSF) through a HCECs monolayer was compared with that of DSF through an excised rabbit cornea. The permeability coefficients of DSF through a HCECs monolayer and excised rabbit cornea were 29.5 +/- 4.8 x 10(-6) cm/s and 34.7 +/- 5.2 x 10(-6) cm/s, respectively. Diethyldithiocarbamate (DDC) had high permeability through HLECs monolayer with a permeability coefficient of 44.6 +/- 7.1 x 10(-6) cm/s. The cytotoxicity of DDC against HLECs was investigated using the trypan blue exclusion test. For a DDC concentration of 5 mmol/l, more than 85% cells were viable. DH3a1 mRNA was expressed in cultured HLECs. The expression of aldehyde dehydrogenase 3a1 (ALDH3a1), which may be be responsible for DSF-DDC conversion, was detected using RT-PCR and agarose gels electrophoresis. These results demonstrate that the permeability of DSF can be detected and intra-ocular drug action may be predicted using the cultured HCEC and HLEC monolayers as model.

Quantitative Modeling and Simulation in PMDA: A Japanese Regulatory Perspective.

In Japan in October 2016, the Pharmaceuticals and Medical Devices Agency (PMDA) began to receive electronic data in new drug applications (NDAs). These electronic data are useful to conduct regulatory assessment of sponsors' submissions and contribute to the PMDA's research. In this article, we summarize the number of submissions of quantitative modeling and simulation (M&S) documents in NDAs in Japan, and we describe our current thinking and activities about quantitative M&S in PMDA.

Clinical evaluation of acute phase nystagmus associated with cerebellar lesions.

OBJECTIVES: To determine the characteristics of acute phase nystagmus in patients with cerebellar lesions, and to identify a useful indicator for differentiating central lesions from peripheral lesions. METHODS: Acute phase nystagmus and the appearance of neurological symptoms were retrospectively investigated in 11 patients with cerebellar stroke. RESULTS: At the initial visit, there were no patients with vertical nystagmus, direction-changing gaze evoked nystagmus or pure rotatory nystagmus. There were four cases with no nystagmus and seven cases with horizontal nystagmus at the initial visit. There were no neurological symptoms, except for vertigo and hearing loss, in any cases at the initial visit. The direction and type of nystagmus changed with time, and neurological symptoms other than vertigo appeared subsequently to admission. CONCLUSION: It is important to observe the changes in nystagmus and other neurological findings for the differential diagnosis of central lesions.

The inhibition of human factor Xa by plasminogen activator inhibitor type 1 in the presence of calcium ion, and its enhancement by heparin and vitronectin.

Plasminogen activator inhibitor type 1 (PAI-1), a member of serine proteinase inhibitor superfamily, is known to inhibit thrombin in the presence of either heparin or vitronectin. We analyzed possible inhibitory activity of PAI-1 on human factor Xa. PAI-1 inhibited factor Xa in the presence of calcium ion (Ca2+), whereas no inhibition was observed in the absence of Ca2+. Half maximal enhancement by Ca2+ was obtained at 0.8 mM. An equimolar complex formation between factor Xa and PAI-1 in the presence of Ca2+ was observed by SDS polyacrylamide gel electrophoresis. Both unfractionated heparin and vitronectin enhanced the inhibition only in the presence of Ca2+. Apparent second-order rate constant (ki) for the inhibition of factor Xa by PAI-1 at 5 mM Ca2+ was 1.6 x 10(4) M-1 s-1, and was enhanced 3-fold by 2 u/ml of heparin (4.6 x 10(4) M-1 s-1) and 10-fold by 100 nM vitronectin (1.6 x 10(5) M-1 s-1), respectively. The interaction between Ca(2+)-bound factor Xa and PAI-1 could be important from the view of PAI-1 neutralization and enhancement of fibrinolysis.

Role of plasminogen system components in focal cerebral ischemic infarction: a gene targeting and gene transfer study in mice.

BACKGROUND: The role of plasminogen system components in focal cerebral ischemic infarction (FCI) was studied in mice deficient in plasminogen (Plg-/-), in tissue or urokinase plasminogen activator (tPA-/- or uPA-/-), or in plasminogen activator inhibitor-1 or alpha2-antiplasmin (PAI-1(-/-) or alpha2-AP-/-). METHODS AND RESULTS: FCI was produced by ligation of the left middle cerebral artery and measured after 24 hours by planimetry of stained brain slices. In control (wild-type) mice, infarct size was 7.6+/-1.1 mm3 (mean+/-SEM), uPA-/- mice had similar infarcts (7.8+/-1.0 mm3, P=NS), tPA-/- mice smaller (2.6+/-0.80 mm3, P<0.0001), PAI-1(-/-) mice larger (16+/-0.52 mm3, P<0.0001), and Plg-/- mice larger (12+/-1.2 mm3, P=0.037) infarcts. alpha2-AP-/- mice had smaller infarcts (2. 2+/-1.1 mm3, P<0.0001 versus wild-type), which increased to 13+/-2.5 mm3 (P<0.005 versus alpha2-AP-/-) after intravenous injection of human alpha2-AP. Injection into alpha2-AP-/- mice of Fab fragments of affinospecific rabbit IgG against human alpha2-AP, after injection of 200 microg human alpha2-AP, reduced FCI from 11+/-1.5 to 5.1+/-1.1 mm3 (P=0.004). CONCLUSIONS: Plg system components affect FCI at 2 different levels: (1) reduction of tPA activity (tPA gene inactivation) reduces whereas its augmentation (PAI-1 gene inactivation) increases infarct size, and (2) reduction of Plg activity (Plg gene inactivation or alpha2-AP injection) increases whereas its augmentation (alpha2-AP gene inactivation or alpha2-AP neutralization) reduces infarct size. Inhibition of alpha2-AP may constitute a potential avenue to treatment of FCI.