A risk as an infection route: Nasal colonization of methicillin-resistant Staphylococcus aureus USA300 clone among contact sport athletes in Japan.

Panton-Valentine leukocidin (PVL)-positive USA300 clone is a highly pathogenic and global epidemic community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) clone. Athletes are particularly vulnerable to CA-MRSA infection because of the frequency of skin trauma, close-contact situations, and sharing of equipment that is customary in the athletic setting. We experienced a case of Japanese collegiate football player with septic pulmonary emboli secondary to infectious iliofemoral deep venous thrombosis caused by the USA300 clone. Here, we screened the nasal carriage of USA300 clone colonization among asymptomatic teammate of the patient to elucidate the infection route. Among 69 nasal samples, CA-MRSA strains were found in 5.8% (four samples). Molecular epidemiological analyses showed that three of the CA-MRSA strains were USA300 clone. Furthermore, pulsed-field gel electrophoresis revealed that all nasal USA300 clones showed 100% identity with the USA300 clone isolated from their teammate with critical infection. Our findings indicate that nasal colonization of the PVL-positive CA-MRSA, especially USA300 clone, pose a threat among contact sport athletes in Japan likewise other countries. An immediate infection control strategy for contact sport athletes is necessary to prevent outbreaks of PVL-positive CA-MRSA infections.

Phase I/II trial of combination of temozolomide chemotherapy and immunotherapy with fusions of dendritic and glioma cells in patients with glioblastoma.

BACKGROUND: This trial was designed to evaluate the safety and clinical responses to a combination of temozolomide (TMZ) chemotherapy and immunotherapy with fusions of DCs and glioma cells in patients with glioblastoma (GBM). METHOD: GBM patients were assigned to two groups: a group of recurrent GBMs after failing TMZ-chemotherapy against the initially diagnosed glioma (Group-R) or a group of newly diagnosed GBMs (Group-N). Autologous cultured glioma cells obtained from surgical specimens were fused with autologous DCs using polyethylene glycol. The fusion cells (FC) were inoculated intradermally in the cervical region. Toxicity, progression-free survival (PFS), and overall survival (OS) of this trial were evaluated. Expressions of WT-1, gp-100, and MAGE-A3, recognized as chemoresistance-associated peptides (CAP), were confirmed by immunohistochemistry of paraffin-embedded tumor samples. Patient's PBMCs of pre- and post-vaccination were evaluated by tetramer and ELISPOT assays. RESULTS: FC-immunotherapy was well tolerated in all patients. Medians of PFS and OS of Group-R (n = 10) were 10.3 and 18.0 months, and those of Group-N (n = 22) were 18.3 and 30.5 months, respectively. Up-regulation and/or cytoplasmic accumulation of CAPs was observed in the recurrent tumors of Group-R patients compared with their initially excised tumors. Specific immune responses against CAPs were observed in the tetramer and ELISPOT assays. CONCLUSIONS: The combination of TMZ-treatment leading to up-regulation and/or cytoplasmic accumulation of CAPs, with FC-immunotherapy as a means of producing specific immunity against CAPs, may safely induce anti-tumor effects in patients with GBM.

Procalcitonin levels predict to identify bacterial strains in blood cultures of septic patients.

BACKGROUND: We examined whether the values obtained from principal component analysis (PCA) on laboratory tests can be used to predict bacterial infections and identify bacterial strains in blood culture (BC). METHOD: This study is a single-center retrospective analysis of 315 patients suspected of having sepsis. We applied PCA on procalcitonin (PCT) and laboratory test biomarkers, namely, platelet (PLT), white blood cell, and C-reactive protein (CRP) as well as BC. RESULTS: Principal component analysis showed that PCT, CRP, and PLT contributions to component 1 were associated with bacterial infection. The number of patients who had BC-negative results, gram-positive cocci (GPC), and gram-negative rods (GNRs) were 124, 28, and 19, respectively. The mean value of component 1 in GNR-positive patients was 1.58±1.41 and was significantly higher than that in GPC-positive patients (0.28±0.87; P<.0001). Furthermore, the mean values of component 1 in both GNR- and GPC-positive patients were significantly higher than that in BC-negative patients (-0.31±0.65; P<.0001 and P<.002, respectively). One certain range showing higher value more than 2.00 for component 1 and -1.00 for component 2 only included GNR-positive patients. There were no BC-positive patients who showed less than -1.00 for component 1. CONCLUSION: The present results obtained by PCA on laboratory tests involving PCT, PLT, white blood cell, and CRP suggest the potential of PCA-obtained values to not only predict bloodstream infection but also identify bacterial strains. This provides some clinical significance in the management of sepsis in acute care.

[Burns].

Burns extending deep into the skin and those affecting a wide surface area trigger various responses in the body and pose a serious threat to life. Therefore, the degree of severity needs to be determined accurately, and appropriate transfusion and local management should be provided accordingly. Systematic and meticulous management that considers not just the risk of death but also functional prognosis is essential from the early stage of burn injuries. Such management requires comprehensive care by a medical team concerning infections, nutrition and rehabilitation. This article outlines the current status of intensive care for severe burns.

[A Case of Foix-Alajouanine Syndrome due to Perimedullary AVF: Remission and Aggravation Mechanism Considered by an Open Surgical Biopsy and Intraoperative ICG Angiography].

Foix-Alajouanine syndrome (FAS), also known as congestive myelopathy due to spinal vascular malformations, presents with paraplegia, sensory disturbance of lower limbs, and dysfunction of the bladder and rectum. Although FAS is characterized by a subacute onset of neurological symptoms that may wax and wane over a few years, the progression mechanism remains unclear. We report a case of FAS due to an angiographically occult arteriovenous fistula (AVF) that was diagnosed by an open surgical biopsy and intraoperative indocyanine green (ICG) angiography. The patient was a 74-year-old female who presented with a one-year history of gradually progressive gait disturbance, weakness, and decreased sensation in her legs associated with bladder and rectum dysfunction. MRI showed intramedullary T1 hypointensity, T2 hyperintensity at level Th4-12, and intramedullary enhancing with a Gd-DTPA lesion at level Th8-12. A true-FISP image of the MRI revealed an abnormal tortuous vessel in the dorsal spinal subarachnoid space, but digital subtraction angiography of the spine at the C1-L5 level showed no abnormality. The patient also underwent Th8-12 laminectomy for open biopsy. ICG angiography revealed blood flow stagnation in an abnormally enlarged posterior spinal vein. Histological findings indicated necrotizing myelopathy and stenosis with wall thickening of the posterior spinal vein. We hypothesized that the progression mechanism in the present case may have resulted from histological changes in the draining veins of an AVF. Intraoperative ICG angiography may be a valuable method, not only for diagnosing AVFs but also for determining the existence and pathological characteristics of FAS.

Prediction of blood culture results by measuring procalcitonin levels and other inflammatory biomarkers.

BACKGROUND: It would be helpful if we could predict positive or negative blood culture results. This study considered the usefulness of measuring procalcitonin (PCT) level and standard clinical biomarkers such as white blood cell (WBC) count, C-reactive protein (CRP) level, and platelet (PLT) count to predict blood culture results. METHOD: We retrospectively analyzed the data from 422 specimens collected at our emergency center within the preceding 36 consecutive months. Primary component analysis (PCA) was used for detecting the degree of the relational contribution of each of the 4 biomarkers to the blood culture results. RESULTS: Procalcitonin alone (cut-off value, 0.5 ng/mL) yielded a positive blood culture rate of 34.0%. Procalcitonin plus 3 biomarkers (WBC, CRP, and PLT) analyzed by PCA yielded 45.9% or 35.3% when a case was in the first or fourth quadrant, which was significantly higher than cases in the second or third quadrant. Primary component analysis also revealed that positive blood culture results were mainly affected by primary component 1, to which PCT and PLT (not WBC or CRP) predominantly contribute. CONCLUSION: Although it is difficult to predict blood culture results, even using 4 biomarkers analyzed by PCA, our new finding that blood culture results are affected not by WBC and CRP, but mainly by PCT and PLT, might help explain the mechanism of sepsis.

[Transient charles bonnet syndrome after excision of a right occipital meningioma: a case report].

Charles Bonnet syndrome is a condition characterized by visual hallucinations. These simple or complex visual hallucinations are more common in elderly individuals with impaired peripheral vision. The current report describes a case of transient Charles Bonnet syndrome appearing after the removal of a meningioma. The patient was a 61-year-old man who already had impaired visual acuity due to diabetic retinopathy. Brain MRI revealed a cystic tumor severely compressing the right occipital lobe. Starting on day 2 postoperatively, the patient was troubled by recurring visual hallucinations involving people, flowers, pictures, and familiar settings(the train and a coffee shop). These continued for 3.5 months. This period roughly coincided with the time for the occipital lobe to recover from the compression caused by the tumor, a fact that was confirmed by several MRI scans. ¹²³I-IMP SPECT performed 1 month after the surgical operation showed an area of hypoperfusion in the right parieto-occipital lobe. Based on the patient's clinical course and MRI findings, the mechanism of onset of visual hallucinations in this patient was put forward. The release of pressure in the brain by tumor removal and subsequent recovery changed the blood flow to the brain. This triggered visual hallucinations in the patient, who was already predisposed to developing Charles Bonnet syndrome because of diabetic retinopathy. This case is interesting since it indicates that central neurological factors, as well as visual deficits, may induce the appearance of visual hallucinations in Charles Bonnet syndrome.

Clinical efficacy and safety of arbekacin sulfate in patients with MRSA sepsis or pneumonia: a multi-institutional study.

Arbekacin (ABK) is an aminoglycoside and widely used in Japan for treatment of patients infected with methicillin-resistant Staphylococcus aureus (MRSA). Although, ABK has concentration-dependent antibacterial activity, the peak serum concentration (C (peak)) of ABK has not yet been fully investigated as an indicator of the efficacy of ABK. The present study was conducted in patients admitted to hospitals affiliated with the ABK Dose Finding Study Group, between October 2008 and June 2011, who had pneumonia or sepsis, the cause of which was identified or suspected to be MRSA. The initial target C (peak) was set at 15-20 µg/mL and therapeutic drug monitoring was conducted. Then the relationship between serum concentration and efficacy/safety of ABK was prospectively examined to obtain sufficient clinical efficacy. In total, 89 patients from 11 clinical sites in Japan were enrolled and 29 of these patients were subjected to efficacy analysis. The mean initial dose and C (peak) were 306.9 mg/day and 16.2 µg/mL, respectively. The efficacy rate was 95 % (19/20 patients) at 5-6 mg/kg or higher, 87.5 % (7/8) for sepsis and 90.5 % (19/21) for pneumonia, and the overall efficacy rate was 89.7 % (26/29). There was no increase in the incidence of adverse events. In conclusion, we recommend the initial dose of ABK at 5-6 mg/kg or higher and the dosage regimen should be adjusted to achieve C (peak) at 10-15 µg/mL or higher in the treatment of patients with pneumonia or sepsis caused by MRSA. This strategy would surely achieve low incidence of adverse events while obtaining high clinical efficacy.

Neoechinulin a imparts resistance to acute nitrosative stress in PC12 cells: a potential link of an elevated cellular reserve capacity for pyridine nucleotide redox turnover with cytoprotection.

Treatment of PC12 cells with fungus-derived alkaloid neoechinulin A for more than 12 h renders the cells resistant to subsequent superoxide (O2⁻)/nitric oxide (NO) insults derived from 3-morpholinosydnonimine (SIN-1). However, the underlying mechanism(s) remains largely unclear. To elucidate the mechanism(s), we assessed the specificity of the cytoprotection afforded by neoechinulin A treatment using other cytocidal stressors and also clarified the resulting cellular alterations, focusing on the antioxidant and metabolic enzymes systems. Neoechinulin A treatment for more than 12 h endowed PC12 cells with significant resistance to transient NO toxicity, but not persistent NO toxicity, bolus H2O2 toxicity, or oxidative insult from the redox cycling quinone menadione. Cellular antioxidant system profiling revealed no substantial potentiation of the activity of any antioxidant enzyme in lysate from the neoechinulin A-treated cells excluding glutathione (GSH) content, which was significantly decreased (>50%), resulting in a proportional compromise in the thiol-reducing activity of the intact cells. In addition, no differences were observed in the activity for any nicotinamide adenine dinucleotide (phosphate) reduced form (NAD(P)H)-generating enzyme, steady-state NAD(P)H/nicotinamide adenine dinucleotide (phosphate) oxidized form (NAD(P)⁺) ratios, or the levels of total NAD(P)H. Nevertheless, the neoechinulin A-treated intact cells exhibited increased NAD(P)H redox turnover when driven by extracellular tetrazolium. The structurally inactive analog preechinulin failed to protect cells against NO toxicity or induce these alterations, suggesting their link with the cytoprotective mechanism. These results suggest that neoechinulin A, despite disabling the GSH defense system, confers cytoprotection against nitrosative stresses by elevating the cellular reserve capacity for NAD(P)H generation, which could offset crippling of energy-supplying systems due to nitrosative stress.

Surgical treatment of a sylvian-middle fossa dural arteriovenous fistula draining into the basal vein of rosenthal with frontotemporal craniotomy.

A 61-year-old man presented with left lower quadrianopsia caused by cerebral infarction in the right occpital lobe. Cerebral angiography revealed occlusion of right transverse sinus and Sylvian-middle fossa dural arteriovenous fistula (d-AVF) draining into the Sylvian vein and dilation of basal vein of Rosenthal. Surgical operation with right frontotemporal craniotomy was carried out to obliterate the fistula point and resection of the dura mater containing vasculature networks. Histologically, the thickening of walls of dural arteries and veins lacking internal elastica lamina were observed. Interestingly, the dura mater involving d-AVF was hyalinized and lacked collagen fibers, resembling local hypoxia and suggesting the possible role of dural hypoxia with pathogenesis of d-AVF. The present case indicates that open surgery can be effective for Sylvian-middle fossa d-AVF for the purpose of obliteration of fistula point and resection of the dura for histopathologic analyses.

Multistage indocyanine green videoangiography for the convexity dural arteriovenous fistula with angiographically occult pial fistula.

Recently, intraoperative indocyanine green (ICG) videoangiography has become a common technique for treating cerebrovascular diseases. We report a case of dural arteriovenous fistula (AVF) treated with direct surgery using intraoperative ICG videoangiography. A 41-year-old man with right hemiplegia caused by a left subcortical hemorrhage was transferred to our hospital. Digital subtraction angiography (DSA) revealed a left convexity parasagittal dural AVF. Surgical resection of the dural AVF was performed using step-by-step ICG videoangiography 4 times in each dissection procedure, which precisely delineated the structure of the dural AVF. After a circular incision of the dura around the fistular point, repeated ICG videoangiography identified the residual fistula between the pial artery from the middle cerebral artery and the draining vein. Complete disappearance of the AVF was confirmed by ICG videoangiography after this pial fistula was removed. Postoperative DSA revealed no residual AVF. Accurate detection of all fistular points and complete resection, including the dura mater and pial vessels, are necessary to avoid rebleeding caused by the residual dural AVF due to incomplete obliteration of the fistular points. Intraoperative ICG videoangiography could provide information on angiographically occult vascular malformation, such as pial fistulas, that cannot be detected by preoperative DSA. Our findings suggest that multistage intraoperative ICG videoangiography can be quite useful for complete resection of a dural AVF with angiographically occult pial fistula.

Nonconvulsive Status Epilepticus Caused by Cerebrospinal Fluid Dissemination of a Salivary Duct Carcinoma: A Case Report.

Salivary duct carcinoma (SDC) is a rare and highly aggressive salivary gland tumor with rapid growth, distant metastasis, and a high recurrence rate. Moreover, the parotid gland is the most common site with a poor prognosis. A lower frequency of distance metastasis to the liver, skin, and brain has also been reported, although the lungs, bones, and lymph nodes are the most common sites of SDC metastasis. We report a case of nonconvulsive status epilepticus (NCSE) in a 73-year-old male comatose patient having SDC of the parotid gland with an unusual metastasis to the skin and brain diagnosed by frequent cerebrospinal fluid examinations. Meningeal carcinomatosis usually has a poor prognosis, and NCSE is a reversible cause of altered mentation. Clinicians should know the unique set of epilepsy etiologies in patients with malignant tumors.

Neoechinulin a impedes the progression of rotenone-induced cytotoxicity in PC12 cells.

Neoechinulin A, an indole alkaloid from marine fungi, can protect PC12 cells from the cytotoxicity of 1-methyl-4-phenylpyridinium (MPP(+)), a Parkinson disease-inducing neurotoxin, by ameliorating downstream events resulting from mitochondrial complex I inactivation. However, the cytoprotective mechanisms remained unclear. In this study, by using rotenone, another parkinsonian-inducing neurotoxin targeting mitochondrial complex I, we investigated the cytoprotective mechanism of neoechinulin A. Rotenone-induced cell death was associated with accelerated glucose consumption, and excess glucose supplementation in the culture medium almost completely suppressed cell death, suggesting that glucose deficiency in the medium is critical for triggering cell death in this model. Co-treatment with neoechinulin A, but not neoechinulin A pre-treatment before rotenone exposure, significantly impeded cell death by rotenone. Although the presence of neoechinulin A did not affect the accelerated glycolytic turnover in rotenone-treated cells, it paradoxically decreased ATP levels in the cells, suggesting increased ATP consumption. Although the link between the decreased ATP levels and cytoprotection is not clear at present, it suggests that neoechinulin A may ameliorate rotenone toxicity by activating a cytoprotective machinery that requires ATP.

Identification of a persistent primitive trigeminal artery following the transposition technique for trigeminal neuralgia: a case report.

A patient who presented with trigeminal neuralgia associated with a persistent primitive trigeminal artery (PPTA) is presented. A 62-year-old woman suffering from right orbital pain was admitted to the hospital. Medical treatment for three months was ineffective, and her neuralgia had deteriorated and gradually spread in the maxillary division. Magnetic resonance imaging demonstrated the flow void signal attached to the right trigeminal nerve. Thus, microvascular decompression was performed. The superior cerebellar artery was the responsible artery, and it was transposed to decompress the trigeminal nerve. After this manoeuvre, an artery was identified running parallel to the trigeminal nerve toward Meckel's cave. The artery, which turned out to be a PPTA, communicated with the basilar artery. The PPTA was carefully observed, and it was found not to be the artery causing the neuralgia because it did not compress the nerve at surgical observation. No additional procedure between the PPTA and the trigeminal nerve was performed. The patient's symptom improved dramatically following surgery, and her postoperative course was uneventful. Postoperative three-dimensional computed tomography showed the PPTA. The findings in the present case suggest that transposition of the responsible artery effectively decompresses the root entry zone and assists in determining whether the PPTA is affecting the trigeminal nerve.

Psychiatric disorders of the combination of levetiracetam either with lacosamide or perampanel: a retrospective cohort study.

Background The number of patients with epilepsy receiving perampanel or lacosamide as an add-on treatment following levetiracetam treatment has increased. Although levetiracetam causes psychiatric disorders, it is unclear whether they occur with the combined use of these antiepileptic drugs. Objective To determine the frequency of psychiatric disorders in patients received lacosamide or perampanel in combination with levetiracetam. Setting A single-center retrospective cohort study. Method Patients who received levetiracetam + lacosamide or levetiracetam + perampanel were selected. Medical records from the start of combination therapy contained characteristics of patients and the incidence of psychiatric disorders. Main outcome measure The frequency of psychiatric disorders, the time to onset, dose reduction or discontinuation following psychiatric disorders, and the clinical course following disorder onset. Results Forty-four patients used levetiracetam + lacosamide and 50 used levetiracetam + perampanel. The incidence of psychiatric disorders was significantly lower (p < 0.001) with levetiracetam + lacosamide (6.8%) than with levetiracetam + perampanel (44%). The incidence of affect lability was significantly higher with levetiracetam + perampanel than with levetiracetam + lacosamide (p = 0.018). The time to the onset of psychiatric disorders was within 1 month of dose initiation or increase in one case (33.3%) with levetiracetam + lacosamide and 16 cases (72.7%) with levetiracetam + perampanel. There was no significant difference in clinical characteristics and antiepileptic drug dosages owing to the presence or absence of psychiatric disorders. Conclusion As the frequency of psychiatric disorders was higher with levetiracetam + perampanel therapy, levetiracetam + lacosamide may be preferable. These disorders tended to develop within 1 month of therapy and were not dose-dependent. Antiepileptic drugs should be cautiously prescribed to avoid psychiatric disorders.

The carboxy-terminal fragment of alpha(1A) calcium channel preferentially aggregates in the cytoplasm of human spinocerebellar ataxia type 6 Purkinje cells.

Spinocerebellar ataxia type 6 (SCA6) is an autosomal dominant neurodegenerative disease caused by a small polyglutamine (polyQ) expansion (control: 4-20Q; SCA6: 20-33Q) in the carboxyl(C)-terminal cytoplasmic domain of the alpha(1A) voltage-dependent calcium channel (Ca(v)2.1). Although a 75-85-kDa Ca(v)2.1 C-terminal fragment (CTF) is toxic in cultured cells, its existence in human brains and its role in SCA6 pathogenesis remains unknown. Here, we investigated whether the small polyQ expansion alters the expression pattern and intracellular distribution of Ca(v)2.1 in human SCA6 brains. New antibodies against the Ca(v)2.1 C-terminus were used in immunoblotting and immunohistochemistry. In the cerebella of six control individuals, the CTF was detected in sucrose- and SDS-soluble cytosolic fractions; in the cerebella of two SCA6 patients, it was additionally detected in SDS-insoluble cytosolic and sucrose-soluble nuclear fractions. In contrast, however, the CTF was not detected either in the nuclear fraction or in the SDS-insoluble cytosolic fraction of SCA6 extracerebellar tissues, indicating that the CTF being insoluble in the cytoplasm or mislocalized to the nucleus only in the SCA6 cerebellum. Immunohistochemistry revealed abundant aggregates in cell bodies and dendrites of SCA6 Purkinje cells (seven patients) but not in controls (n = 6). Recombinant CTF with a small polyQ expansion (rCTF-Q28) aggregated in cultured PC12 cells, but neither rCTF-Q13 (normal-length polyQ) nor full-length Ca(v)2.1 with Q28 did. We conclude that SCA6 pathogenesis may be associated with the CTF, normally found in the cytoplasm, being aggregated in the cytoplasm and additionally distributed in the nucleus.

Application of an automated cardiopulmonary resuscitation device for kidney transplantation from uncontrolled donation after cardiac death donors in the emergency department.

Vital-organ transplantation has become acceptable as the treatment of choice for end-stage organ failure. If the patient, facing the end of life, wishes to donate organs after cardiac arrest (CA), donation after cardiac death (DCD) is increasingly important for the realization of the patient's desires after CA. In Japan, kidney transplantation from uncontrolled DCD donors, who are identified in modified Maastricht categories II or V, is one of the critical factors in expanding the donor pool. However, according to the forensic code for post-mortems and the requirement of legal consent for transplantation, the time required to meet all procedural requirements has sometimes prohibited organ procurement from uncontrolled DCD donors. We have therefore attempted to use an automated cardiopulmonary resuscitation (CPR) device and maintain arterial pressure for uncontrolled DCD donors during all interim procedures after sudden CA. Comparing kidneys procured from standard DCD donors (n = 10) and uncontrolled DCD donors (n = 4), significant differences were seen in warm ischemic time (WIT), defined as the time from CA to initiation of cooling in situ. However, our early experience showed good tolerance and viability of uncontrolled DCD kidneys. Immediate availability of an automated CPR device might provide a bridge to kidney procurement from uncontrolled DCD donors.

First Report of Fatal Infection Caused by Community-acquired Methicillin-resistant Staphylococcus aureus USA300 Clone in a Collegiate Athlete.

Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) is prevalent around the world and is a causative agent of skin and soft tissue infections in healthy individuals. Particularly, Panton-Valentine leukocidin (PVL)-positive CA-MRSA strains occasionally cause life-threatening infections, such as septic pulmonary emboli (SPE) and infectious endocarditis. However, severe infections caused by PVL-positive CA-MRSA strains have rarely been reported in Japan. For the first time, this study reports the case of a 20-year-old Japanese college athlete with life-threatening PVL-positive CA-MRSA USA300 clone infection, including sepsis, SPE, and skin and soft tissue infections with iliofemoral deep venous thrombosis.

SIN-1 cytotoxicity to PC12 cells is mediated by thiol-sensitive short-lived substances generated through SIN-1 decomposition in culture medium.

As a generator of peroxynitrite (ONOO(-)), 3-morpholinosydnonimine (SIN-1) is widely used in the study of oxidative/nitrosative stress in cultured cells, although controversy exists regarding active species responsible for cytotoxicity. In this study, we report that unstable thiol-sensitive substances, generated from the reaction of SIN-1 with components in culture medium, play a crucial role in SIN-1 cytotoxicity in PC12 cells. Exposure of cells to culture medium obtained after almost complete SIN-1 decomposition at 37 degrees C for 2h demonstrated almost the same degree of cytotoxicity as did fresh SIN-1. The cytotoxicity of SIN-1-decomposed medium largely depended on serum, decayed with time, and could be completely abolished by the addition of thiols. Degradation of synthetic ONOO(-) in the culture medium did not reproduce the unstable cytotoxicity. The presence of superoxide dismutase (SOD) during SIN-1 decomposition prevented the formation of the cytotoxic substances, whereas SOD had no protection against the cytotoxicity itself, suggesting a crucial role of simultaneously generated superoxide and nitric oxide in the formation of the toxicants, but not in their cytotoxic action. The cytotoxicity of fresh SIN-1 is dramatically suppressed in a basal medium (Hanks balanced salt), suggesting that the cytotoxicity of fresh SIN-1 also requires components of culture medium. These results suggest that SIN-1 cytotoxicity in PC12 cells is mediated via the generation of cytotoxic substances in the medium during its decomposition.