[Improvement of oncologic results of transurethral resection in the treatment of non-muscle-invasive bladder cancer].

AIM: To evaluate the efficiency of additional methods of intraoperative control during transurethral resection (TUR) for the treatment of non-muscle invasive bladder cancer. MATERIAL AND METHODS: A total of 138 patients (92 men and 46 women) with non-muscle-invasive bladder cancer (Ta-T1N0M0) were treated in the urological clinic of Kazan State Medical University. The median age was 59 years. In 28 patients TUR was performed as monotherapy, in 28 patients TUR with photodynamic therapy (PDD) was done and other 26 patients undergone TUR under dynamic transurethral ultrasound control. In 29 patients, TUR was combined with a single intravesical instillation of a chemotherapy drug, and in 27 patients, TUR was combined with long-term intravesical chemotherapy. The frequency and type of relapses was evaluated depending on the treatment method during five-year follow-up period. Analysis of postoperative complications and their severity was performed according to the Clavien-Dindo classification. Statistical analysis was performed using the Statistica 7.0 and Microsoft Excel 2003 software package. Survival was assessed using the Kaplan-Meier method. Differences in survival between groups were determined using a log-rang test. RESULTS: The total 5-years recurrence rate in the group 1 was 60.71% (n=17). There were 6 recurrences in the resection area (21.43%) and 8 recurrences outside the resection area (28.57%). The progression rate was 10.71% (n=3). In the group 2, the overall recurrence rate was 25% (n=7), including 2 (7.14%) and 4 (14.29%) recurrences in and outside resection area, respectively. The progression rate was 3.57% (n=1). In the group 3, where TUR was performed in combination with transurethral ultrasound, 7 recurrences were diagnosed over a five-year period (26.92%), including 1 recurrence in the resection area (3.84%) and 6 recurrences in other parts of bladder (23.07%). There was no progression of bladder cancer. In the group of patients who received a single intravenous chemotherapy after TUR, there were no significant differences with the group of patients where TUR was performed as monotherapy. The total number of recurrences was 55.16% (n=16). There were 4 recurrences in the resection area (13.79%) and 9 recurrences in other parts of bladder (31.03%), as well as 3 case of disease progression (10.34%). At the same time, in the group of patients where prolonged course of adjuvant intravesical chemotherapy was performed, a significant decrease in the recurrences rate in the resection area (7.4%; n=2) and progression (3.7%, n=1) was found. The number of recurrences outside the resection area was comparable with the group 1 (22.22%; n=6). CONCLUSIONS: According to our data, we recommend to perform TUR in combination with PDD and transurethral ultrasound in order to improve the oncological results. Long-term intravesical chemotherapy is an effective alternative in case of inability to use additional intraoperative control and it should be included in the treatment scheme of patients with a high risk of recurrence.

Susac syndrome: clinical characteristics and treatment in 29 new cases.

BACKGROUND AND PURPOSE: There are few clinical studies on the attempted treatments and outcomes in patients with Susac syndrome (SS) (retinocochleocerebral vasculopathy). METHODS: A retrospective review was performed of all patients presenting with SS at the Mayo Clinic in Rochester, Minnesota, USA (1 January 1998-1 October 2011). RESULTS: There were 29 cases of SS (24 women, mean age at presentation, 35 years; range, 19-65; full triad of brain, eye, and ear involvement, n = 16; mean follow-up time, 29 months). Thirty CSF analyses were performed in 27 cases (mean protein 130 mg/dl, range 35-268; mean cell count 14, range 1-86). MRI of the brain showed corpus callosal involvement (79%), T2-weighted hyperintensities (93%), and gadolinium enhancement (50%). Average lowest modified Rankin Scale score was 2.5 (median 2, range 0-5). Most patients (93%) received immunosuppressive treatment, with a mean time to treatment of 2 months following symptomatic onset. Treatments included intravenous methylprednisolone or dexamethasone (n = 23), oral corticosteroids (n = 24), plasma exchange (PLEX) (n = 9), intravenous immunoglobulin (IVIg) (n = 15), cyclophosphamide (n = 6), mycophenolate mofetil (n = 5), azathioprine (n = 2), and rituximab (n = 1). Most patients also received an antiplatelet agent (n = 21). Improvement or stabilization was noted in eight of 11 cases treated with IVIg in the acute period (three experienced at least partial deterioration) and eight of nine cases of PLEX treatment (one lost to follow up). CONCLUSIONS: Susac syndrome may be severe, disabling, and protracted in some patients. PLEX may be an adjunct or alternative therapy for patients who do not experience symptomatic improvement following steroid treatment.

Role of MAPK in chronic cerebral vasospasm.

This study was undertaken to investigate the role of p44/42 MAPK in a dog double hemorrhage model of subarachnoid hemorrhage (SAH), and whether MEK inhibitors can alter the degree of SAH-induced vasoconstriction. The diameter of the basilar artery, which was compared with day 0 angiogram, decreased gradually in a time-dependent manner from day 3 (80%), day 5 (68%) through day 7 (53.5%). The level of MAPK (p44/42) immunoprecipitation peaked on day 3 and remained enhanced through day 7 (P < 0.05). MEK inhibitor PD98059 significantly reduced p44/42 MAPK immunoprecipitation and significantly reversed vasospasm and increased residual diameter to 79.0% on day 7. These results demonstrated that p44/42 MAPK kinase is involved in the pathogenesis of cerebral vasospasm. The MEK inhibitor PD98059 might be useful in the treatment of vasospasm.

Oxyhemoglobin produces apoptosis and necrosis in cultured smooth muscle cells.

Confluent rat aortic smooth muscle cells were treated with OxyHb in a concentration- and time-dependent manner. A high concentration of OxyHb (100 microM) within 24 h decreased cell density. DNA analysis showed a smear pattern characteristic of cell necrosis. Transmission electron microscopy demonstrated disintegration of the cell membrane and destruction of cell organelles. Western blotting using PARP antibody revealed that 116 kDa PARP was not cleaved to 85 kDa, an apoptosis-related fragment. On the contrary, a low concentration of OxyHb (10 microM) produced apoptotic cell death at 72 h that was supported by DNA analysis and TUNEL staining. These results demonstrated that a high level of OxyHb induced necrosis within 24 h and a low concentration of OxyHb produced apoptosis after 72 h in cultured smooth muscle cells. Morphological alterations induced by OxyHb might contribute to the vascular wall changes in the cerebral arteries following subarachnoid hemorrhage (SAH).

Relaxant effect of U0126 in hemolysate-, oxyhemoglobin-, and bloody cerebrospinal fluid-induced contraction in rabbit basilar artery.

BACKGROUND AND PURPOSE: It has been suggested that mitogen-activated protein kinase (MAPK) is involved in cerebral vasospasm after subarachnoid hemorrhage. The present study was undertaken to explore the inhibitory effect of U0126, a novel MAPK inhibitor, in the contraction of the rabbit basilar artery by 3 spasmogens: hemolysate, oxyhemoglobin, and bloody cerebrospinal fluid (CSF) from patients with vasospasm. METHODS: The contraction and relaxation of rabbit basilar arteries were measured by isometric tension. MAPK immunoprecipitation was assessed by Western blot analysis. RESULTS: (1) Pretreatment of the rabbit basilar arteries with U0126 reduced contractions to hemolysate, oxyhemoglobin, or bloody CSF applied subsequently. (2) In the absence of endothelial cells, U0126 produced an inhibitory effect similar to the contractions induced by hemolysate, oxyhemoglobin, or bloody CSF. (3) U0126 relaxed the sustained contraction induced by hemolysate, oxyhemoglobin, or bloody CSF. (4) Hemolysate, oxyhemoglobin, and bloody CSF enhanced MAPK immunoprecipitation. (5) U0126 reduced MAPK immunoprecipitation induced by hemolysate, oxyhemoglobin, and bloody CSF. (6) Hemolysate, oxyhemoglobin, and bloody CSF significantly increased MAPK activity in the rabbit basilar artery. (7) U0126 abolished the effect of hemolysate, oxyhemoglobin, or bloody CSF on MAPK activation. CONCLUSIONS: This study demonstrated a role of MAPK in the contraction of rabbit basilar arteries by hemolysate, oxyhemoglobin, and bloody CSF. MAPK inhibitor U0126 may be useful in the treatment of cerebral vasospasm.

Prevention of vasospasm in penetrating arteries with MAPK inhibitors in dog double-hemorrhage model.

BACKGROUND: Vasospasm in the penetrating arteries contributes to ischemic neurological deficit. It may be as important as angiographic vasospasm because it would explain the discrepancies between angiographic vasospasm and clinical symptoms in some patients. It may also underlie the different effects of vasodilators. The present study examined this hypothesis by looking at the effect of the inhibitors of mitogen-activated protein kinase (MAPK) on vasospasm of the penetrating arteries. METHODS: Twenty-two adult mongrel dogs of either sex were used for the dog double-hemorrhage model. The dogs were randomly divided into four groups: control-hemorrhage, vehicle-treated, PD98059-treated, and U0126-treated groups. The drug injections were started on Day 3 after the first subarachnoid hemorrhage (SAH). The clinical status of the dogs was studied, based on their activity, appetite, and focal neurological symptoms. On Day 7, all the dogs were sacrificed, and the penetrating arteries from the brain stem were prepared for transmission electron microscopy. RESULTS: (1) Severe vasospasm developed in the basilar arteries in the SAH-without-treatment group (control), in the DMSO-treated group (DMSO), and in the U0126 treatment group with mean reduction of the basilar artery diameter of 46.57%, 49.3%, and 39.6%, respectively. In the PD98059-treatment group only a mild vasospasm was observed and the mean reduction of the basilar artery diameter was 18.9%. (2) All the dogs in the control SAH and vehicle-treated groups developed severe angiographic and clinical vasospasm. The penetrating arteries were contracted, and the endothelial and smooth muscle cells were dystrophic. (3) The dogs in the U0126-treated group developed severe angiographic, but not clinical, vasospasm. The penetrating arteries were not contracted, and the endothelial and smooth muscle cells were not dystrophic. (4) The dogs in the PD98059 group developed mild angiographic vasospasm. No dog developed clinical symptoms that could be attributed to vasospasm. In morphological studies, the penetrating arteries were slightly contracted, but the cells were not dystrophic. CONCLUSIONS: Vasospasm of the penetrating arteries, but not angiographic vasospasm, is consistent with the clinical symptoms and signs of vasospasm. MAPK may be important in maintaining vasospasm of both major and penetrating cerebral arteries. The correlation of the improvement in the clinical score with the reduction of vasospasm in the penetrating arteries demonstrated an important role of penetrating arteries in the morbidity and mortality caused by SAH.

Morphological changes of cerebral penetrating arteries in a canine double hemorrhage model.

BACKGROUND: Morphological presentations of cerebral vasospasm, such as dystrophy and desquamation of endothelial cells, corrugation of the internal elastic layer, and necrotic changes in smooth muscle cells, are well defined in large cerebral arteries. This study was undertaken to examine pathological changes in cerebral penetrating arteries in a canine double hemorrhage model. METHODS: Eighteen mongrel dogs were subjected to an autologous arterial blood (0.4 mL/kg) injection into the cisterna magna on day 0 and day 2 after withdrawal of an equivalent amount of cerebrospinal fluid. Angiogram was performed on day 0 before the blood injection and on the day the dogs were sacrificed. The dogs were divided into four groups: control (day 0) (n = 4), hemorrhage and sacrificed on day 3 (n = 4), day 5 (n = 5), and day 7 (n = 5). The penetrating arteries were removed and found to be spastic on days 3, 5, and 7, but not in the control group. RESULTS: Endothelial dystrophy and partial desquamation were recorded in all dogs sacrificed on days 5 and 7. Condensation of chromatin, blebbing of the membrane, and condensation of cytoplasm were identified in many endothelial cells, features that are consistent with apoptosis. The morphological changes in the penetrating arteries were more pronounced on days 5 and 7. CONCLUSIONS: Vasospasm occurred in cerebral penetrating arteries in a canine double hemorrhage model. The morphological change in penetrating arteries, especially apoptosis in endothelial cells, is consistent with an early phase of vasospasm. Vasospasm in a penetrating artery may contribute to the cerebral ischemia that occurs during vasospasm.

Mechanism of ATP-induced [Ca(2+)](i) mobilization in rat basilar smooth muscle cells.

BACKGROUND AND PURPOSE: We have previously reported that extracellular ATP activates P(2u) receptors and increases intracellular free Ca(2+) ([Ca(2+)](i)) by G protein/phospholipase C/inositol 1,4,5-triphosphate pathways in cerebral artery smooth muscle cells. However, the possible contribution of other signaling pathways remains unclear. This study was undertaken to investigate the role of protein tyrosine kinase (PTK) and mitogen-activated protein kinase (MAPK) in mediating ATP-induced Ca(2+) mobilization in rat basilar artery smooth muscle cells (RBASMCs). METHODS: RBASMCs were freshly isolated, and [Ca(2+)](i) was monitored by fura 2 microfluorimetry. MAPK phosphorylation was studied by the Western blot technique. RESULTS: ATP produced a biphasic [Ca(2+)](i) response, which consists of releasing Ca(2+) from internal stores and influx from extracellular space. PTK inhibitors tyrphostin 51 and genistein inhibited [Ca(2+)](i) response to ATP. Tyrphostin A1, an inactive analogue of tyrphostins, failed to reduce the ATP-induced response. MAPK kinase inhibitor PD98059, but not U0126, reduced the ATP-induced [Ca(2+)](i) response. Phosphatidylinositol 3-kinase (PI3-K) tyrosine kinase inhibitor wortmannin, but not janus tyrosine kinase (JAK2) inhibitor AG490, partially inhibited the [Ca(2+)](i) response induced by ATP. In addition, ATP enhanced MAPK phosphorylation in a concentration- and time-dependent manner, and genistein, tyrphostin 51, PD98059, and U0126 inhibited MAPK phosphorylation. CONCLUSIONS: Extracellular ATP produced [Ca(2+)](i) elevation and MAPK phosphorylation in RBASMCs, and the effect was regulated by PTK. The role of MAPK in ATP-induced [Ca(2+)](i) elevation is not clear. PI3-K tyrosine kinase and JAK2 tyrosine kinase may not play an important role in the ATP-induced [Ca(2+)](i) response in RBASMCs.

Apoptosis of endothelial cells in vessels affected by cerebral vasospasm.

BACKGROUND: Cerebral vasospasm after subarachnoid hemorrhage is a prolonged contraction that leads to cerebral ischemia or infarction. Morphological studies of cerebral arteries during vasospasm have shown extensive necrosis of smooth-muscle cells and desquamation and dystrophy of endothelial cells. The mechanism of cellular death is unknown. METHODS: We report an observation of apoptotic changes in the cerebral arteries of a patient who died after suffering severe cerebral vasospasm caused by aneurysmal rupture. Subarachnoid hemorrhage and cerebral vasospasm were confirmed by computed tomography scanning and angiogram. Histological and immunohistological examinations for apoptosis were performed in cerebral arteries. For control, the arteries from another patient, who died of trauma without head injury, were used. RESULTS: Corrugation of the internal elastic lamina and increased amounts of connective tissue was demonstrated by light microscopy. Apoptotic changes, characterized by condensation of chromatin of the nucleus and detachment from the basal membrane, were found on transmission electron microscopy in endothelial cells. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling reaction revealed positive staining of the nuclei of the endothelial cells. CONCLUSIONS: This study demonstrates that apoptosis occurred in the cerebral arteries in a patient who died of cerebral vasospasm. The possible role of apoptosis in cerebral vasospasm is discussed.

Effect of endothelin receptor antagonists on non-muscle matrix compaction in a cell culture vasospasm model.

Endothelin-1 (ET-1), a potent vascular smooth muscle constrictor, is one of the possible spasmogens in cerebral vasospasm. However, the role of ET-1 in non-muscle compaction (another aspect of the pathogenesis of cerebral vasospasm) has not been reported. This study was undertaken to demonstrate the effect of ET-1, as well as erythrocyte lysate and bloody cerebrospinal fluid (CSF), on fibroblast populated collagen lattice (FPCL) compaction. Human dermal fibroblasts were used to form FPCL. The concentration-dependent effect of ET-1 was examined in the absence and presence of an ETA receptor antagonist (BQ-485), or an ETB receptor antagonist (BQ-788), or both. FPCL compaction was determined by measuring reduction of areas over five days following treatment. To compare the effect of ET-1 on lattice compaction, erythrocyte lysate and bloody CSF obtained from a cerebral vasospasm patient were also tested. We found that ET-1 increased FPCL compaction in a concentration-dependent (but not time-dependent) manner. Erythrocyte lysate produced the strongest compaction, however, without time-dependence. Bloody CSF promoted FPCL compaction in a time-dependent fashion. Compaction induced by ET-1 was inhibited by BQ-485 but not by BQ-788. We concluded that ET-1 promotes FPCL compaction by activation of ETA receptors. Other components in bloody CSF or erythrocytes may also contribute to FPCL compaction.

Risk factors for the development of post-traumatic cerebral vasospasm.

BACKGROUND: Post-traumatic vasospasm is a well-recognized sequela of head injury. The risk factors associated with post-traumatic vasospasm have not been well defined. We studied 119 consecutive patients with head injury to determine the risk factors for post-traumatic vasospasm. METHODS: Twenty-nine (27.1%) patients were excluded from the study because of poor insonation (n = 12) or a hospital stay of less than 72 hours (n = 17). Seventy (77.8%) of 90 patients suffered severe head injury. Sixteen (17.8%) patients sustained moderate head injury and four (4.4%) patients sustained mild head injury. All patients were monitored with transcranial Doppler (TCD) ultrasonography daily. RESULTS: Post-traumatic vasospasm was detected in 32 (35.6%) of 90 patients. Among these patients, 29 (90.6%) had severe head injury, and three (9.4%) had moderate head injury. None of the patients with mild head injury suffered post-traumatic vasospasm. In most cases, the onset of post-traumatic vasospasm began on the fifth day and lasted 1 to 9 days. In 8 (25%) patients, post-traumatic vasospasm began within the first three days of the head injury. Among 32 patients with post-traumatic vasospasm, 10 (31.2%) patients had mild vasospasm, 20 (65.5%) had moderate vasospasm, and 2 (6.3%) had severe post-traumatic vasospasm. Clinical deterioration was documented in two (2.5%) patients. CONCLUSIONS: Development of post-traumatic vasospasm correlated only with severe subarachnoid hemorrhage on initial computed tomographic scan. There was an increased incidence of post-traumatic vasospasm in patients with epidural hematomas, subdural hematomas, and intracerebral hemorrhages. The Glasgow Coma Scale (GCS) score on admission was inversely related to the development of post-traumatic vasospasm. In most cases, the period of vasospasm was short and clinical deterioration was rare. Probably, two varieties of post-traumatic vasospasm exist, one that lasts a shorter time and does not correlate with the presence of SAH, and a second that correlates with the presence of SAH, lasts longer, and resembles aneurysmal vasospasm.

Mechanism of endothelin-1-induced contraction in rabbit basilar artery.

BACKGROUND AND PURPOSE: Endothelin-1 (ET-1) is suggested to be a major cause of cerebral vasospasm after subarachnoid hemorrhage. However, the mechanism of ET-1-induced contraction in cerebral arteries remains unclear. This study was undertaken to demonstrate the possible role of protein tyrosine kinase (PTK), mitogen-activated protein kinase (MAPK), and protein kinase C (PKC) in ET-1-induced contraction. METHODS: PD-98059, damnacanthal, wortmannin, AG-490, genistein, calphostin C, and staurosporine were used to inhibit, or relax, the ET-1-induced contraction of basilar artery, studied with an isometric tension system. Immunoprecipitation of MAPK in ET-1-stimultated rings of basilar artery without or with the above inhibitors was studied with Western blot. RESULTS: (1) ET-1 produced concentration-dependent contraction and MAPK immunoprecipitation in rabbit basilar artery by activation of ET(A) but not ET(B) receptors. (2) MAPK inhibitors PD-98059 and U-0126 produced dose-dependent inhibition of ET-1-induced contraction. (3) The Src tyrosine kinase inhibitor damnacanthal, the phosphatidylinositol-3 kinase inhibitor wortmannin, and the Janus tyrosine kinase(2) inhibitor AG-490 abolished ET-1-induced contraction. (4) The PKC inhibitor staurosporine but not calphostin C abolished ET-1-induced contraction, and the PTK inhibitor genistein partially reduced ET-1-induced contraction. (5) In arteries precontracted by ET-1, PD-98059, U-0126, wortmannin, AG-490, genistein, and staurosporine produced concentration-dependent relaxation. (6) ET-1 induced a biphasic and time-dependent MAPK immunoprecipitation. (7) PD-98059, U-0126, genistein, AG-490, and damnacanthal, but not staurosporine or wortmannin, abolished the effect of ET-1 on MAPK immunoreactivity. CONCLUSIONS: This study demonstrated that MAPK may be involved in ET-1-induced contraction in rabbit basilar artery. MAPK is downstream of PTK, Src, and Janus tyrosine kinase pathways but may not be downstream of phosphatidylinositol-3 kinase pathways. The possible involvement of PKC in ET-1-induced contraction requires further investigation. Inhibition of these pathways may offer alternative treatment for ET-1-induced contraction and cerebral vasospasm.

Mitogen-activated protein kinase plays an important role in hemolysate-induced contraction in rabbit basilar artery.

OBJECT: Mitogen-activated protein kinase (MAPK) is an important signaling factor in the vascular proliferation and contraction, the two features of cerebral vasospasm following subarachnoid hemorrhage. We studied the possible involvement of MAPK in hemolysate-induced signal transduction and contraction in rabbit basilar artery. METHODS: Isometric tension was used to record the contractile response of rabbit basilar artery to hemolysate. Western blots using antibodies for MAPK were conducted. 1) Hemolysate produced a concentration-dependent contraction of rabbit basilar artery. Pre-incubation of arteries with MAPK kinase inhibitor PD-98059 markedly reduced the contraction induced by hemolysate. PD-98059 also relaxed, in a concentration-dependent fashion, the sustained contraction induced by hemolysate (10%). 2) Hemolysate produced a time-dependent elevation of MAPK immunoreactivity in Western blot in rabbit basilar artery. MAPK was enhanced 3 min after hemolysate exposure and the effect reached maximum at 5 min. The immunoreactivity of MAPK decayed slowly with time, but the level of MAPK was still higher than the basal level even at two hours after exposure to hemolysate. 3) Pre-incubation of arteries with MAPK kinase inhibitor PD-98059 abolished the effect of hemolysate on MAPK immunoreactivity. CONCLUSION: Hemolysate produced contraction of rabbit basilar artery possibly by activation of MAPK. MAPK inhibitors may be useful in the treatment of cerebral vasospasm.

Morphological presentation of posttraumatic vasospasm.

Posttraumatic vasospasm is a well-recognized sequela of head injury. The risk factors associated with posttraumatic vasospasm have not been well defined. We studied 119 consecutive patients with head injury to determine the risk factors for posttraumatic vasospasm. Posttraumatic vasospasm was detected in 32 (35.6%) of 90 patients. Among these patients, 29 (90.6%) had severe head injury and 3 (9.4%) had moderate head injury. None of the patients with mild head injury suffered posttraumatic vasospasm. In most cases, the onset of posttraumatic vasospasm began on the fifth day and lasted 1 to 9 days. In 8 (25%) patients, posttraumatic vasospasm began within the first three days of the head injury. Clinical deterioration was documented in two (2.5%) patients. Morphologically, posttraumatic vasospasm resembled features of aneurysmal vasospasm. We found increased corrugation of the internal elastic lamina and increased amounts of connective tissue in the subendothelial layer. These findings showed that posttraumatic vasospasm, although clinically more mild, demonstrated the same morphological changes as did aneurysmal vasospasm.

Oxyhemoglobin produces necrosis in cultured smooth muscle cells.

OBJECT: Myonecrosis in the tunica media, which is defined morphologically, is one of the most striking alterations in the cerebral arterial wall following subarachnoid hemorrhage (SAH). In this study, oxyhemoglobin (OxyHb) was added to cultured rat aortic smooth muscle cells to determine the pattern of cell death by morphological and biochemical techniques. METHODS: Confluent rat aortic smooth muscle cells were treated with OxyHb in a concentration- and time-dependent manner. Cell density was assayed by counting the number of cells that attached to the culture dishes after exposed to OxyHb. To identify cell death pattern, DNA analysis, electron microscopy, and Western blotting using poly (ADP-ribose) polymerase (PARP) antibody were performed. CONCLUSIONS: OxyHb decreased cell density in a concentration- and time-dependent manner. DNA analysis showed a smear pattern characteristic of cell necrosis. Transmission electron microscopy demonstrated disintegration of cell membrane and destruction of cell organelles. No apoptotic changes, such as condensation of chromatin or apoptotic bodies were observed. Western blotting using PARP antibody revealed that 116 kDa PARP was not cleaved to 85 kDa, an apoptosis-related fragment. These results demonstrated morphologically and biochemically that OxyHb induced necrosis, not apoptosis, in cultured smooth muscle cells.

Posttraumatic cerebral vasospasm: clinical and morphological presentations.

Head injury is one of the leading causes of morbidity and mortality in the young population. Many factors complicate head injury and worsen an outcome. One of these factors is posttraumatic cerebral vasospasm. We studied 75 patients admitted to the University of Mississippi Medical Center with head injury. Their ages ranged from 14 to 67 years (mean 30 years, SD 11.63). Eighty percent of the patients were men, and 20% were women. Of these patients, 53 (70.6%) suffered severe blunt trauma, and 4 patients suffered gunshot wounds to the head. Four patients had mild head injury, and 14 had moderate head injury. Posttraumatic vasospasm was detected in 24 (32%) patients. Among these patients, 19 (79.2%) had severe closed head injury, 3 patients had moderate head injury, and 2 suffered gunshot wounds. The severity of the patient's respective condition was correlated with the development of posttraumatic cerebral vasospasm: 50% of the patients with Glasgow Coma Scale (GCS) 3-4 developed PTV, and only 30% with GCS 9-11, and none of the patients with GCS > 12 developed PTV. Overall, posttraumatic vasospasm started earlier and had a shorter course than did aneurysmal vasospasm. Morphologically, posttraumatic vasospasm resembled the features of aneurysmal vasospasm. We found increased corrugation of the internal elastic lamina and increased amounts of connective tissue in the subendothelial layer. These findings show that posttraumatic vasospasm, although clinically more mild, demonstrates the same morphological changes as aneurysmal vasospasm.

Oxyhemoglobin-induced apoptosis in cultured endothelial cells.

OBJECT: Oxyhemoglobin (OxyHb) is one of the most important spasmogens for cerebral vasospasm that follows aneurysmal subarachnoid hemorrhage. The cytotoxic effect of OxyHb has been documented in endothelial and smooth-muscle cells; however, the pattern of cell death--necrosis or apoptosis--as the final stage of cell damage has not been demonstrated. This study was undertaken to determine if OxyHb induces apoptotic changes in cultured bovine aortic endothelial cells. METHODS: Confluent bovine aortic endothelial cells were treated with OxyHb in a concentration- and time-dependent manner. Cell density was assayed by counting the number of cells that attached to culture dishes after exposure to OxyHb. To identify apoptotic changes, the investigators used three specific methods: DNA fragmentation (electrophoreses), the apoptotic body (transmission electron microscopy), and cleavage of poly (adenosine diphosphate ribose) polymerase (PARP [Western blotting]). CONCLUSIONS: Oxyhemoglobin decreased cell density in a concentration- and time-dependent manner. Analysis of DNA showed a pattern of internucleosomal cleavage characteristic of apoptosis (DNA ladder). Transmission electron microscopy demonstrated condensation of nuclei and apoptotic bodies in OxyHb-treated endothelial cells. Western blotting with the PARP antibody revealed that the 116-kD PARP was cleaved to the 85-kD apoptosis-related fragment. These results for the first time demonstrated that the OxyHb induces apoptosis in cultured endothelial cells.