Effect of SMTP-7 on Cisplatin-Induced Nephrotoxicity in Mice.

SMTP-7, a fungal metabolite, is reported to have a high degree of availability for the ischemia-reperfusion (IR)-induced acute kidney injury (AKI) model. Cisplatin, a widely used anticancer drug, has serious side effects, such as AKI. Hence, we aimed to examine the effect of SMTP-7 on cisplatin-induced AKI in this study. Significant increases in blood urea nitrogen (BUN) and serum creatinine (Scr) were observed at 72 h after the intravenous infusion of cisplatin (20 mg/kg). Histologically, necrosis and dilatation (hyaline casts) as well as regeneration were observed in proximal tubules. SMTP-7 inhibited the elevation on BUN and Scr caused by cisplatin dose dependently. The efficacy of SMTP-7 was notable when the drug was administered on the day after cisplatin treatment, whereas the repeated administration of the drug did not result in an enhanced efficacy. Moreover, 10 mg/kg of SMTP-7 considerably ameliorated tubular necrosis and dilatation. The cisplatin treatment also caused an up-regulation of tumor necrosis factor-α (TNF-α) mRNA expression prior to the elevation of the levels of BUN and Scr. Administration of SMTP-7 (10 mg/kg) at 24 h after the cisplatin infusion alleviated the up-regulation of TNF-α mRNA expression. These findings suggest that SMTP-7 exhibits a renoprotective effect against cisplatin infusion based on the inhibition of the expression of pro-inflammatory cytokines such as TNF-α and may be expected a new effective drug for the treatment of cisplatin-induced AKI.

Acetic acid treatment causes renal inflammation and chronic kidney disease in mice.

We established a novel mouse model of chronic kidney disease (CKD) using acetic acid and compared it with the 5/6-nephrectomized mouse model. In our novel model, significant increases were observed in blood biochemical values and urinary parameters. Moreover, a decrease in creatinine clearance (Ccr) was observed. This model also demonstrated a higher survival rate than the 5/6-nephrectomized model. Observed histological changes in our model included cell infiltration in the renal interstitium, tubular dilation, regenerated tubules, and glomerulosclerosis. Inflammation of the renal interstitium was particularly remarkable. TNF-α, IL-1ß, and ICAM-1 mRNA expression were up-regulated prior to elevation of mean blood pressure and prior to changes in blood biochemical values and urinary parameters. Up-regulation of TGF-ß mRNA and down-regulation of nephrin mRNA were also observed at 12 weeks after acetic acid treatment. However, no correlation between the progression of CKD and the decrease in renal blood flow was observed. Finally, repeated losartan administration attenuated the effects of acetic acid-induced renal injury. Our findings suggest that chronic kidney conditions associated with this model may be triggered by interstitial inflammation. Moreover, we suggest that this model is useful for understanding the pathophysiological mechanisms of CKD, and for evaluating the effects of therapeutic agents.

Evaluation of the effects of a new series of SMTPs in the acetic acid-induced embolic cerebral infarct mouse model.

We reported previously that Stachybotrys microspora triprenyl phenol-7 (SMTP-7) showed potential thrombolytic, anti-inflammatory and anti-oxidant effects that account for its excellent pharmacological activity such as having a wider therapeutic time window than tissue plasminogen activator (t-PA) and a significant protection against hemorrhage. The aim of the present study was to evaluate and compare the effect of a new series of SMTPs in the acetic acid-induced embolic cerebral infarct mouse model. Thrombotic occlusion was produced in mice by inducing the transfer of acetic acid-induced thrombi from the right common carotid artery into the brain. SMTPs were evaluated by their effect on reducing infarct area, neurological score and edema. Furthermore, plasmin formation, anti-inflammatory and anti-oxidant activities were assessed by fibrin zymography, measuring pro-inflammatory gene expression, and thiobarbituric acid reactive substances (TBARS) assay, respectively. Treatment with either SMTP-22 or SMTP-43 (10mg/kg), which have similar plasmin formation, anti-inflammatory and anti-oxidant activities to SMTP-7, resulted in reduced infarct area, neurological score and edema. Coexistence of all these three activities appears to be important for the treatment of embolic infarction because SMTP-6, SMTP-25, and SMTP-44D (10mg/kg), which are each missing at least one of the three functions, were not as effective. Therefore, these results indicate that SMTP-22 and SMTP-43 have potential as medicinal compounds for the treatment of embolic cerebral infarction.

[A Case of Stage IV Breast Cancer with Long-Term Survival by Chemotherapy Developing Pulmonary Tumor Thrombotic Microangiopathy during the Treatment Course].

Pulmonary tumor thrombotic microangiopathy(PTTM)caused by pulmonary artery microscopic tumor emboli and fibrocellular and/or fibromuscular proliferation leads to progressive pulmonary hypertension and respiratory failure.The prognosis is extremely poor and most patients die shortly after onset.We report a patient with Stage IV breast cancer and long-term survival who developed PTTM during chemotherapy treatment.A 63-year-old woman with multiple metastases in her cerebellum, bone, lung, and lymph node after left breast conserving surgery started to experience dyspnea and malaise 7 years after the surgery.Two months later, she was urgently admitted to hospital because of respiratory failure and was diagnosed with pulmonary hypertension.However, pulmonary thrombosis and tumor thrombus were not observed.We clinically diagnosed her with PTTM and administered chemotherapy in addition to treatment for pulmonary hypertension.Her medical condition improved gradually and she survived for the subsequent 2 years.When observing progressive hypoxia and pulmonary hypertension without obvious pulmonary embolism findings on imaging, PTTM should be considered.Early diagnosis and immediate induction of chemotherapy for primary disease can improve the survival of patients with PTTM.

Change in soluble epoxide hydrolase (sEH) during cisplatin-induced acute renal failure in mice.

Cisplatin is one of the most effective chemotherapeutic agents against various types of cancers; however, it is also associated with nephrotoxicity. Recently, it was reported that inflammatory mechanisms play a key role in the development of nephrotoxicity. Epoxyeicosatrienoic acids (EETs) have an anti-inflammatory effect and are metabolized by soluble epoxide hydrolase (sEH: encoded by EPHX2 gene). Here, we determined the change in sEH activity and EPHX2 expression in renal tissue associated with the development of cisplatin-induced nephrotoxicity. Cisplatin administration decreased hydrolase activity accompanied by down-regulation of sEH and EPHX2 expression. The down-regulation occurred prior to the elevation of blood urea nitrogen (BUN) and tumor necrosis factor-α (TNF-α) gene expression or at treatment with low dose cisplatin. In addition, a negative correlation was found between EPHX2 expression and renal thiobarbituric acid reactive substance (TBARS), and edaravone, a radical scavenger, administration did not down-regulate expression of this gene. The results of this study suggest that cisplatin decreased sEH activity through the down-regulation of sEH and EPHX2 expression, and this down-regulation was involved in a negative feedback loop to protect renal tissue from further damage. Thus, sEH is a potential therapeutic target of cisplatin-induced nephrotoxicity.

Two types of overcontraction are involved in intrarenal artery dysfunction in type II diabetic mouse.

Contractile responses in small intrarenal arteries are associated with diabetic nephropathy. However, the mechanisms that induce and maintain altered small vessel contraction are not clearly understood. To further understand intrarenal artery dysfunction in diabetes, phenylephrine (PE)-induced force development was assessed in the intrarenal artery [interlobar artery (ILA)] in control (lean) and type II diabetic (ob/ob) mice. PE-induced dose-dependent force development in the ILA was significantly greater in ob/ob mice than in lean mice (592.8 ± 5.2 and 770.1 ± 12.1 µ/mm tissue, respectively, following administration of 30 µM PE, n = 5). Under high-glucose conditions (twice the normal concentration of glucose), PE-induced force development in the ILA was only enhanced in ob/ob mice (946.0 ± 18.2 µN/mm tissue; n = 5). ILA dysfunction reduces blood flow to the glomerulus and may induce diabetic nephropathy. Basal overcontraction of the ILA in ob/ob mice under normal-glucose conditions was reduced by pretreatment with rottlerin, a calcium-independent protein kinase C (PKCδ) inhibitor. Total PKC activity was also reduced by rottlerin. Under high-glucose conditions, the enhanced ILA contraction in diabetic mice was suppressed by rho A and rho kinase inhibitors. Our results indicate two types of ILA dysfunction in diabetes, as follows: 1) a basal increase in PE-induced contraction under normal-glucose conditions, and 2) extracellular glucose-dependent enhancement of PE-induced contraction. We believe that these dysfunctions are mediated by the activation of the PKCδ and rho A-rho kinase pathways, respectively.

Altered gene expression in an embolic stroke model after thrombolysis with tissue plasminogen activator and Stachybotrys microspora triprenyl phenol-7.

The present study compares gene expression and infarct area in a mouse model of embolic stroke after thrombolysis with t-PA and SMTP-7. Embolic occlusion was induced by transfer of acetic acid-induced embolus into the brain. t-PA or SMTP-7 was administered 3 h after embolization. Changes in gene expression were evaluated using microarray and RT-PCR analysis. To determine the involvement of reactive oxygen species in the response to t-PA, the free radical scavenger edaravone was infused immediately before t-PA administration. The expressions of 459 genes involved in the inflammatory response, cell-to-cell signaling, cell movement, and inflammatory disease were altered by embolic occlusion. Twenty-two of those genes were upregulated after t-PA but not SMTP-7 administration. Differences between the t-PA- and SMTP-7-treated groups in the expression of genes including the proinflammatory genes Il6, Stat3, S100a8, and Mmp9 were confirmed with RT-PCR. Edaravone ameliorated the overexpression of these genes. Our data demonstrate differences in gene expression following treatment with SMTP-7 or t-PA that likely explain the difference in therapeutic time windows of the two drugs. ROS are involved in the overexpression of proinflammatory genes. The wide therapeutic time window may be achieved through an anti-oxidative effect and inhibition of proinflammatory gene overexpression.

SMTP-7, a novel small-molecule thrombolytic for ischemic stroke: a study in rodents and primates.

SMTP-7 (Stachybotrys microspora triprenyl phenol-7), a small molecule that promotes plasminogen activation through the modulation of plasminogen conformation, has excellent therapeutic activity against cerebral infarction in several rodent models. Detailed evaluations of SMTP-7 in a primate stroke model are needed for effective, safe drug development. Here we evaluated SMTP-7 in a monkey photochemical-induced thrombotic middle cerebral artery (MCA) occlusion model (n=6), in which MCA occlusion was followed by recanalization/reocclusion. SMTP-7 (10 mg/kg, intravenous infusion) significantly increased the postinfusion MCA recanalization rate (32.5-fold, P=0.043) and ameliorated the post-24-h neurologic deficit (by 29%, P=0.02), cerebral infarct (by 46%, P=0.033), and cerebral hemorrhage (by 51%, P=0.013) compared with the vehicle control animals. In normal monkeys, SMTP-7 did not affect general physiologic or hemostatic variables, including coagulation and platelet parameters. Investigations in rodent models of transient and permanent focal cerebral ischemia, as well as arterial thrombosis and bleeding tests, suggest a role for SMTP-7's regulated profibrinolytic action and neuroprotective properties in the monkey MCA occlusion model. In conclusion, SMTP-7 is effective in treating thrombotic stroke in monkeys. SMTP-7 is thus a promising candidate for the development of alternative therapy for ischemic stroke.

Lysophosphatidic acid induces shear stress-dependent contraction in mouse aortic strip in situ.

We previously reported that lysophosphatidic acid (LPA) regulates Ca²âº influx of fluid flow in stimulated endothelial cells and that LPA and shear stress showed increment and suppressive effects on phenylephrine-induced vasoconstriction and acetylcholine-induced vasodilatation, respectively. However, a vasoconstrictive effect of LPA alone in the presence of shear stress was not found. The present study examined the effect of LPA alone in the presence of shear stress on Ca²âº responses in endothelial and smooth muscle cells and contraction in mouse aortic strip using real-time 2-photon laser scanning microscopy and a custom-made parallel-plate flow chamber. Application of micromolar LPA and high shear stress elicited movement of endothelial cells after Ca²âº responses. The endothelial cells moved along the major axis of smooth muscle cells, a direction that was identical to that found during vasoconstriction evoked by the application of phenylephrine. The frequency of Ca²âº oscillations in smooth muscle cells was highest according to endothelial movement. Vasoconstriction evoked by LPA and shear stress was significantly reduced by the application of a thromboxane A2 receptor antagonist, a cyclooxygenase inhibitor, and a thromboxane synthase inhibitor. These results suggest that micromolar LPA and high shear stress elicit vasoconstriction that is caused by Ca²âº-dependent contraction in medial smooth muscle cells. Thromboxane A2 may be involved in that response.

Adiponectin enhances calcium dependency of mouse bladder contraction mediated by protein kinase Cα expression.

Adiponectin is an adipose tissue-secreted protein and is a multifunctional adipocytokine. However, the association of adiponectin with bladder contraction has not been investigated. In this study, the adiponectin-sense transgenic mouse (Adip-Sen mouse; age, 16-24 weeks; male) and age-matched controls (C57Bl mouse) were studied. The Adip-Sen mouse showed a significant increase in plasma adiponectin levels (56.2%; P < 0.01), compared with those in the C57Bl mouse, without affecting other lipid parameters. Isometric force development in bladder smooth muscle tissues were detected using an organ-bath system. Although carbachol (CCh)-induced (0.1-100 µM) time- and dose-dependent contractions in Adip-Sen mouse bladder were slightly enhanced, compared with those in the C57Bl mouse during a low range (0.3-1.0 µM) of CCh, differences could not be detected with other CCh concentrations. However, the reduction in contraction under Ca(2+)-replaced conditions was significantly different between Adip-Sen and C57Bl mice (94.1 and 66.3% of normal contraction, respectively; n = 5). A parameter of Ca(2+) sensitivity, the relation between intracellular Ca(2+) concentration and contraction, was increased in the Adip-Sen mouse, compared with that in the C57B1 mouse. This Ca(2+) dependency in the Adip-Sen mouse was reduced by a protein kinase C (PKC) inhibitor, but not by a Rho kinase inhibitor. Expression of the calcium-dependent isoform of PKC, PKCα, was increased in the Adip-Sen mouse bladder, and CCh-induced phosphorylation of PKCα was also enhanced, compared with those in the C57Bl mouse. In conclusion, adiponectin is associated with bladder smooth muscle contraction, which involves an increase in Ca(2+) dependency of contraction mediated by PKCα expression.

Impact of chronic total coronary occlusion on microvascular reperfusion in patients with a first anterior ST-segment elevation myocardial infarction.

BACKGROUND: We investigated an impact of the presence of chronic total coronary occlusion (CTO) in a non-infarct related coronary artery on microvascular reperfusion in patients with a first anterior ST-segment elevation myocardial infarction (STEMI) who underwent percutaneous coronary intervention (PCI). METHODS: In accordance with the presence or absence of CTO in a non-infarct related coronary artery, we analyzed Thrombolysis in Myocardial Infarction myocardial perfusion (TMP) grade on a scale of 0 to 3, with higher scores indicating better perfusion, and ST-segment resolution in sum of lead I, aVL, and V1 through V6 to evaluate microvascular reperfusion in a total of 140 consecutive patients with a first anterior STEMI. RESULTS: We identified CTO in 15 patients (11% of total). The incidence of impaired microvascular reperfusion was greater in patients with CTO vs without CTO, defined as TMP grades 0 or 1 together with <30% ST-segment resolution (33% vs 6%, respectively; P=.0006) and the enzymatic infarct was larger (10304 ± 8060 IU/L vs 6804 ± 4959 IU/L; P=.009). Logistic regression analysis revealed that CTO is closely associated with incidental impaired microvascular reperfusion (odds ratio, 6.801; 95% confidence interval, 1.284-36.209; P=.024). CONCLUSION: The presence of CTO in a non-infarct related coronary artery might confer a considerable disadvantage upon microvascular reperfusion and result in adverse clinical outcomes of PCI for a first anterior STEMI.

Two distinct dysfunctions in diabetic mouse mesenteric artery contraction are caused by changes in the Rho A-Rho kinase signaling pathway.

Diabetic complications are associated with small artery dysfunctions. The objective of this study was to identify differences in endothelial cell-denuded mesenteric artery second branch (mesenteric artery-2) contraction, as a typical small artery, between diabetic and non-diabetic mice. Contractile responses in mesenteric artery-2 were assessed in male type 2 diabetic ob/ob mice aged 16-22 weeks and in age-matched control (Lean) mice. Phenylephrine induced dose-dependent contractions in Lean mice (1126.8 ± 28.6 mN/mm tissue at 10 µM phenylephrine; n=5), which were significantly reduced in ob/ob mice (716.8 ± 40.8 mN/mm at 10 µM phenylephrine; n=5). Exposure to high glucose (HG; twice the normal glucose [NG] concentration) enhanced phenylephrine-induced contraction in Lean (1341.4 ± 15.5 mN/mm; n=5) but not in ob/ob mice. These dysfunctions did not involve α(1)-receptor sensitization or protein kinase activity, although the calcium sensitivity of contraction was decreased in ob/ob mice. The Rho kinase inhibitor Y27632 suppressed the difference between Lean and ob/ob mice under NG conditions, which was accompanied by Rho A inactivation. Under HG conditions, glucose-dependent Rho A activation persisted in ob/ob mice whereas Rho kinase expression was reduced. These data suggest that inactivation of Rho A reduced contractibility under NG conditions, and the lack of glucose dependency is associated with reduced Rho kinase expression.

Spatiotemporal dynamics of intracellular calcium in the middle cerebral artery isolated from stroke-prone spontaneously hypertensive rats.

The spatiotemporal dynamics of intracellular calcium within the middle cerebral artery (MCA) isolated from stroke-prone spontaneously hypertensive rats (SHR-SP) were investigated using real-time confocal laser microscopy. At 3 months of age (prestroke), rhythmical changes in the [Ca(2+)](i) during the tonic phase were found to precede vasomotion following application of 5-HT, but not other stimuli. These responses were not observed at 1 month of age; moreover, the MCA lost both responses post-stroke (5 months of age). When [Ca(2+)](i) was analysed in arteriolar smooth muscle cells, rhythmical changes in [Ca(2+)](i) occurred during the same cycle. Thus, these processes were synchronized. The synchronized rhythmical changes in [Ca(2+)](i) were abolished following application of 100 nM ketanserin and 10 µM nicardipine. Treatment with 60 nM charybdotoxin and 10 µM cyclopiazonic acid also significantly reduced rhythmical elevations in [Ca(2+)](i). In addition, rhythmical changes in [Ca(2+)](i) became unsynchronized following treatment with 100 µM carbenoxolone, a gap junction blocker. Connexin 45 mRNA and protein expression were both elevated in the MCA of SHR-SP. Taken together, these findings suggest that rhythmical changes in [Ca(2+)](i) of the MCA are dependent upon the 5-HT(2) receptor-mediated release of calcium from intracellular stores which, in turn, activates voltage-dependent calcium channels to enable an influx of calcium into smooth muscle cells. Subsequently, charybdotoxin-sensitive potassium channels are activated and provide a negative feedback pathway to regulate [Ca(2+)](i). Moreover, the co-ordinated synchronization of rhythmical changes in [Ca(2+)](i) across smooth muscle cells was found to be dependent upon gap junctions.

Clinical features of early myocardial rupture of acute myocardial infarction.

We assessed the clinical features of patients with myocardial rupture within 48 to 72 hours, defined as early myocardial rupture, after percutaneous coronary intervention (PCI) for ST-segment elevation acute myocardial infarction (STEMI). Six patients (4 men, 66 ± 13 years) with early myocardial rupture were identified from 1252 consecutive patients undergoing PCI for STEMI. We evaluated the degree of microvascular reperfusion using thrombolysis in myocardial infarction (TIMI) myocardial perfusion (TMP) grade and a resolution of sum of ST-segment elevation in a 12-lead electrocardiogram (ECG). Time from PCI to myocardial rupture was 11 ± 7 hours. All patients showed TMP grade 0 or 1 and an increase in sum of ST-segment elevation after PCI (1.9 ± 0.5 vs 2.5 ± 0.7 mV; P = .032), suggesting severely failed reperfusion at the level of microcirculation as the common feature to develop early myocardial rupture after PCI for STEMI.

Neuroprotective mechanisms of SMTP-7 in cerebral infarction model in mice.

Reactive oxygen species (ROS) formation has been found to induce the brain damage following stroke-like events. The aim of the present study was to investigate the effect of Stachybotrys microspora triprenyl phenol-7 (SMTP-7) on the generation of ROS in ischemia-induced cerebral infarction model and in vitro lipid peroxidation. We used immunohistochemistry and real-time reverse-transcription PCR for ex vivo evaluation and thiobarbituric acid-reactive substance reagent assay for in vitro evaluation. We demonstrated that SMTP-7 did not induce enhancement of 4-hydroxynonenal or neutrophil cytosolic factor 2 like t-PA administration at 3 h after ischemia ex vivo and reduce lipid peroxidation in vitro. This compound is the first low molecular weight compound with triplet activities of thrombolytic, anti-inflammatory, and antioxidant activities. We theorized that SMTP-7 is among the pharmacological agents that reduce ROS formation and have been found to limit the extent of brain damage following stroke-like events.

Distinct effects of tissue-type plasminogen activator and SMTP-7 on cerebrovascular inflammation following thrombolytic reperfusion.

BACKGROUND AND PURPOSE: Thrombolysis therapy using tissue-type plasminogen activator (t-PA) is occasionally accompanied by harmful outcomes, including intracerebral hemorrhage. We have reported that Stachybotrys microspora triprenyl phenol-7 (SMTP-7), a candidate thrombolytic drug, has excellent therapeutic effect on cerebral infarction in embolic stroke model in mice; however, little is known regarding whether this agent influences cerebrovascular inflammation following thrombolytic reperfusion. The current study aimed to compare the effects of recombinant t-PA (rt-PA) and SMTP-7 on cerebrovascular inflammation. METHODS: The impact of rt-PA- and SMTP-7-induced thrombolytic reperfusion on leukocyte dynamics was investigated in a photochemically induced thrombotic middle cerebral artery occlusion (tMCAo) model in mice. RESULTS: Both rt-PA and SMTP-7 administration in tMCAo mice (each 10 mg/kg) resulted in thrombolytic reperfusion. The SMTP-7-administered mice showed relatively mild rolling and attachment of leukocytes to the vascular wall in the middle cerebral vein, with weak peroxynitrite reactions and proinflammatory gene expression (IL-1ß, TNF-α, ICAM-1, and VCAM-1); thus, a small infarct volume compared with rt-PA-administered mice. In vitro study suggested that rt-PA at 20 µg/mL, but not SMTP-7 at a similar concentration, promotes cytokine-induced reactive oxygen species generation in cultured endothelial cells; moreover, SMTP-7 suppressed cytokine-induced VCAM-1 induction in the cells and leukocyte/ endothelial cell adhesions. CONCLUSIONS: Relatively mild cerebrovascular inflammation and cerebral infarction in the SMTP-7 mice, compared with in rt-PA mice, is thought to be caused at least in part by direct antioxidative actions of SMTP-7 in ECs.

Shear stress-dependent effects of lysophosphatidic acid on agonist-induced vasomotor responses in rat mesenteric artery.

We have previously shown that lysophosphatidic acid (LPA), a bioactive plasma lysophospholipid, markedly accelerates shear stress-induced Ca2+ responses in cultured vascular endothelial cells (ECs). This study aimed to demonstrate the impact of LPA and luminal shear stress on vasomotor regulation in the isolated rat mesenteric artery (MA) using a videomicroscopic technique. Although the addition of LPA to the perfusate in a concentration range of 0.03-0.3 µM had no significant effect on the basal MA tone, LPA in a similar concentration range led to increased phenylephrine-induced MA contraction and reduced acetylcholine-induced MA relaxation under physiological shear conditions. These vasomodulatory actions of LPA, which vanished upon removal of ECs, were positively dependent on luminal shear stress levels and were markedly inhibited by the LPA receptor antagonist Ki16425, the cyclooxygenase inhibitor indomethacin, and the thromboxane A2 receptor antagonist SQ29548. These data thus suggest that LPA can modify the agonist-induced vasomotor responses in MAs in a shear stress-dependent manner. This effect of LPA was mediated through ECs, the LPA receptor, and cyclooxygenase/thromboxane A2 signaling.

A novel embolic model of cerebral infarction and evaluation of Stachybotrys microspora triprenyl phenol-7 (SMTP-7), a novel fungal triprenyl phenol metabolite.

The aim of the present study was to establish a novel embolic model of cerebral infarction and to evaluate the effect of Stachybotrys microspora triprenyl phenol-7 (SMTP-7), a novel fungal triprenyl phenol metabolite. Thrombotic occlusion was induced by transfer of acetic acid-induced embolus into the brain. The regional cerebral blood flow was measured by a laser Doppler flowmeter to check the ischemic condition. Infarction area was assessed by 2% 2,3,5-triphenyltetrazolium chloride (TTC) staining. Neurological scores were determined by a modified version of the method described by Longa et al. Emboli were accumulated at the temporal or parietal region of the middle cerebral artery. Additionally, we found that this model showed decreased cerebral blood flow and increased infarction area and neurological scores. Treatment with tissue plasminogen activator (t-PA) reduced infarction area and the neurological scores in a dose-dependent manner; moreover, the decreased cerebral blood flow recovered. SMTP-7 also reduced these values. The therapeutic time window of SMTP-7 was longer than that of t-PA. These results indicate that this model may be useful for understanding the pathophysiological mechanisms of cerebral infarction and evaluating the effects of therapeutic agents. Additionally, SMTP-7 is a promising approach to extend the therapeutic time window. Therefore, this novel compound may represent a novel approach for the treatment of cerebral infarction.

A novel finding of a low-molecular-weight compound, SMTP-7, having thrombolytic and anti-inflammatory effects in cerebral infarction of mice.

Tissue plasminogen activator (t-PA) has a short therapeutic time window for administration (3 h) and carries a risk of promoting intracerebral hemorrhage. The aim of the present study was to investigate a therapeutic time window and frequency of hemorrhagic region by treatment with Stachybotrys microspora triprenyl phenol-7 (SMTP-7). Thrombotic occlusion was induced by transfer of acetic acid-induced thrombus at the right common carotid artery into the brain of mice. Infarction area, neurological score, edema percentage, and regional cerebral blood flow (CBF) were determined as the index of the efficacy of SMTP-7. In order to evaluate the mechanism of SMTP-7, plasmin activities and the expressions of interleukin (IL)-1beta, tumor necrosis factor-alpha (TNF-alpha), and IL-6 mRNA were examined. SMTP-7 (0.1, 1, 10 mg/kg) dose dependently reduced infarction area, neurological score, and edema percentage. Additionally, its therapeutic time window was longer than that of t-PA, a high-molecular-weight compound. In addition, little hemorrhagic region was induced by treatment with SMTP-7. SMTP-7 showed plasmin activity in vivo and caused a decreased CBF to recover. Furthermore, the expressions of inflammatory cytokine mRNA (IL-1beta, TNF-alpha, IL-6) were increased by t-PA treatment 3 h after ischemia but were not induced by SMTP-7 treatment. These results indicate that SMTP-7 shows potential thrombolytic and anti-inflammatory effects as well as a wide therapeutic time window and little hemorrhagic region compared with that of t-PA. Therefore, this novel low-molecular-weight compound may represent a novel approach for the treatment of cerebral infarction.