The integrin linked kinase (ILK) induces an invasive phenotype via AP-1 transcription factor-dependent upregulation of matrix metalloproteinase 9 (MMP-9).

Overexpression of Integrin Linked Kinase (ILK) in intestinal and mammary epithelial cells results in a highly invasive phenotype, associated with increased levels of expression of the matrix metalloproteinase MMP-9. This increase was at the transcriptional level as determined by MMP-9 promoter-CAT reporter assays. Mutations in the two AP-1 binding sites within the MMP-9 promoter completely inhibited the reporter activity. We have previously shown that ILK inhibits glycogen synthase kinase-3 (GSK-3) activity. Transient transfection of wild-type GSK-3beta in ILK-overexpressing cells decreased MMP-9 promoter activity and AP-1 activity, indicating that ILK can stimulate MMP-9 expression via GSK-3beta and AP-1 transcription factor. A small molecule inhibitor of the ILK kinase reduced the in vitro invasiveness of ILK-overexpressing cells as well as the invasiveness of several human brain tumor cell lines. Furthermore, both MMP-9 promoter and AP-1 activities were inhibited by the ILK inhibitor. Invasiveness of ILK-overexpressing cells was also reduced by inhibition of MMP-9. These data demonstrate that ILK can induce an invasive phenotype via AP-1-dependent upregulation of MMP-9.

Integrin-linked kinase (ILK): a "hot" therapeutic target.

Integrin-mediated cell adhesion is known to regulate gene expression through the activation of transcription factors. We have recently revealed that these activations are mediated through integrin-linked kinase (ILK). ILK is an ankyrin repeat-containing serine-threonine protein kinase that can interact directly with the cytoplasmic domain of the beta1 and beta3 integrin subunits and whose kinase activity is modulated by cell-extracellular matrix interactions. We have shown that ILK overexpression results in the translocation of beta-catenin to the nucleus, which then forms a complex formation with the lymphoid enhancer binding factor 1 (LEF-1) transcription factor, subsequently activating the transcriptional activity of promoters containing LEF-1 response elements. ILK phosphorylates the glycogen synthase kinase-3 (GSK-3), which inhibits GSK-3 activity. We have demonstrated that ILK stimulates activator protein-1 transcriptional activity through GSK-3 and the subsequent regulation of the c-Jun-DNA interaction. ILK also phosphorylates protein kinase B (PKB/Akt) and stimulates its activity. We have shown that ILK is an upstream effector of the phosphatidylinositol 3-kinase-dependent regulation of PKB/Akt. ILK has been shown to phosphorylate PKB/Akt on Ser-473 in vitro and in vivo. Our results clearly indicate that ILK is a key element in the regulation of integrin signaling as well as growth factor and Wnt signaling pathways. PTEN (phosphatase and tensin homolog detected on chromosome 10) is a tumor suppressor gene located on chromosome 10q23 that encodes a protein and phospholipid phosphatase. It is now estimated that inactivation mutants of PTEN exist in 60% of all forms of solid tumors. Loss of expression or mutational inactivation of PTEN leads to the constitutive activation of PKB/Akt via enhanced phosphorylation of Thr-308 and Ser-473. We have demonstrated that the activity of ILK is constitutively elevated in PTEN mutant cells. A small molecule ILK inhibitor suppresses the phosphorylation of PKB at the Ser-473 but not the Thr-308 site in the PTEN mutant cells. These results indicate that inhibition of ILK may be of significant value in solid tumor therapy.

Cell-extracellular matrix interactions stimulate the AP-1 transcription factor in an integrin-linked kinase- and glycogen synthase kinase 3-dependent manner.

Integrin-mediated interactions of cells with components of the extracellular matrix regulate cell survival, cell proliferation, cell differentiation, and cell migration. Some of these physiological responses are regulated via activation of transcription factors such as activator protein 1 (AP-1). Integrin-linked kinase (ILK) is an ankyrin repeat containing serine-threonine protein kinase whose activity is rapidly and transiently stimulated by cell-fibronectin interactions as well as by insulin stimulation. ILK activates protein kinase B and inhibits the glycogen synthase kinase 3 (GSK-3) activity in a phosphatidylinositol-3-kinase (PI 3-kinase)-dependent manner. We now show that cell adhesion to fibronectin results in a rapid and transient stimulation of AP-1 activity. At the same time, the kinase activity of ILK is stimulated whereas that of GSK-3 is inhibited. This fibronectin-dependent activation of AP-1 activity is inhibited in a dose-dependent manner if the cells are transfected with wild-type GSK-3, and also by inhibitors of PI 3-kinase. Stable or transient overexpression of ILK results in a stimulation of AP-1 activity which is inhibited by cotransfection with wild-type GSK-3 and kinase-deficient ILK. Transient transfection of ILK in HEK-293 cells stimulates complex formation between an AP-1 consensus oligonucleotide and nuclear proteins containing c-jun. The formation of this complex is inhibited by cotransfection with active GSK-3 or kinase-deficient ILK, suggesting that ILK may regulate AP-1 activation by inhibiting GSK-3, which has previously been shown to be a negative regulator of AP-1. In the presence of serum, ILK has no effect on the phosphorylation of Ser-73 in the N-terminal transactivation domain of c-jun. These results demonstrate a novel signaling pathway for the adhesion-mediated stimulation of AP-1 transcriptional activity involving ILK and GSK-3 and the subsequent regulation of the c-jun-DNA interaction.

Left ventricular diastolic function after coronary artery bypass grafting: a correlative study with three different myocardial protection techniques.

BACKGROUND: This study was designed to examine the effect of myocardial protection on diastolic function after cardiac operations. METHODS: Subjects were patients with normal preoperative diastolic function who were scheduled for coronary artery bypass grafting. Group I received anterograde cardioplegia; group II received anterograde and retrograde cardioplegia; and group III was protected with ventricular fibrillation and intermittent aortic crossclamping. Operations were performed with mild hypothermia and ventricular venting through the left superior pulmonary vein in all cases. Left ventricular diastolic function was evaluated with pulsed-wave Doppler transesophageal echocardiography (samples at the mitral valve leaflet: four-chamber view) and left superior pulmonary vein flow velocity. The flow patterns were stored on videotape and sent to an independent investigator for analysis. Left ventricular ejection fraction was calculated with transesophageal echocardiography (short-axis view, two-dimensional and M-mode). RESULTS: Left ventricular diastolic function, as measured by the ratio between the peak velocities during early filling and atrial contraction and by systolic diastolic superior pulmonary venous flow ratio, was significantly impaired in all three groups 5 minutes after discontinuation of cardiopulmonary bypass. At 1 hour after operation, these values had returned to control levels only in group III. There was an increased incidence of supraventricular arrhythmias in group III. There were no significant hemodynamic differences among the three groups. CONCLUSIONS: Left ventricular diastolic function was severely impaired after cardiopulmonary bypass. The degree of impairment depended on the myocardial protection used. The impairment in diastolic function was less when ventricular fibrillation and intermittent aortic crossclamping were used, and greater when anterograde and retrograde cardioplegia were used.

Yeast hormone response element assays detect and characterize GRIP1 coactivator-dependent activation of transcription by thyroid and retinoid nuclear receptors.

The mouse glucocorticoid receptor-interacting protein (GRIP1) is a member of the ERAP160 family of nuclear receptor (NR) coactivators (including SRC-1 and TIF2) which function as bridging proteins between ligand-activated NRs bound to cognate hormone-response elements (HREs) and the transcription initiation apparatus (TIA). Although these coactivators bind to several NRs, studies overexpressing these coactivators with these NRs in mammalian cells have not uniformly observed a corresponding enhancement of ligand-dependent transactivation. Here, we show that GRIP1 interacts in vitro in a ligand-dependent manner with thyroid receptor, retinoic acid receptor, and retinoid X receptor. Additionally, in yeast (Saccharomyces cerevisiae) GRIP1 coactivator protein markedly increased the ability of these full-length class II NRs to transactivate beta-galactosidase reporter genes containing cognate HREs. The magnitude of GRIP1 enhancement of liganded NR homodimer was dependent upon NR subtype and HRE configuration. For most HRE configurations, thyroid receptor and retinoic acid receptor homodimers were essentially unresponsive or very weakly active in the absence of GRIP1, but GRIP1 dramatically restored the ligand-dependent function of these NRs. Although GRIP1 exerted no significant effect on NR homodimers in the absence of their cognate ligands, it increased the transactivation of unliganded NR heterodimers. Whether GRIP1 increased ligand-dependent transactivation of a heterodimer to levels greater than that of the cognate homodimer was determined by HRE configuration and copy number. Compared with the limitations of yeast two-hybrid and mammalian coexpression systems, the yeast HRE-assay systems described in this report facilitated both the detection of putative mammalian NR coactivator function and the elucidation of their mechanisms of transactivational enhancement.

Magnesium and arrhythmias after coronary artery bypass surgery.

Arrhythmias are very common after cardiac surgery and are multifactorial. Magnesium is receiving increased consideration in the management of supraventricular and ventricular arrhythmias. This study was designed to evaluate the role of magnesium in preventing arrhythmias in hypokalemic (K < 3.5 mEq/L) and normokalemic (K > 3.5 mEq/L) patients with normal renal and ventricular function after coronary artery bypass grafting (CABG). One hundred forty patients ranging from 32 to 71 years of age who were scheduled for CABG were studied. They were divided into four groups: group I (control) received no magnesium; group II received 10 mg/kg of magnesium sulfate intravenously before cardiopulmonary bypass (CPB); group III received 10 mg/kg of magnesium soon after CPB; group IV received 10 mg/kg of magnesium before and after CPB. Serum potassium and catecholamine levels, as well as serum and urine magnesium levels, were measured and the incidence and type of arrhythmias were determined. There was a statistically significant difference in the occurrence of arrhythmias between the groups studied. The incidence of arrhythmias was highest in groups I and II and lowest in group IV (12 patients in group I, 14 in group II, 5 in group III; and 1 in group IV). Magnesium levels were higher in group IV than any other group studied after completion of surgery. There was no difference in serum and urine magnesium levels between the hypokalemic and normokalemic patients within each group. Serum magnesium returned to normal in all patients after 48 hours. Therefore, it appears that administration of magnesium during and after cardiac surgery reduces the incidence of arrhythmias in hypokalemic and normokalemic patients.

Direct binding of yeast transcription factor (TFIID) to the ribosomal protein L32 (rpL32) TATA-less promoter sequence.

The ribosomal protein L32 (rpL32) gene transcribed by RNA polymerase II lacks a canonical TATA element, that binds the transcription factor TFIID tau or TBP (TATA binding protein). Instead this promoter contains an element, termed gamma, located at -30 relative to the transcription initiation site. We previously reported that, despite the lack of a canonical TATA element the rpL32 gene utilizes yeast TFIID tau for its transcriptional initiation. Whether TFIID tau participates in rpL32 gene transcription by binding directly to a promoter element or through another protein has not been resolved. These studies reveal that proteins ranging in size from 20-40 kDa binds to the gamma-element. The 40 kDa protein(s) displays strong affinity for the canonical TATA element and may be related or equivalent to TFIID tau. Furthermore, cloned and purified yeast TFIID (TBP) binds directly to the gamma-element implying that the gamma-element directs RNA polymerase II-dependent transcription of the rpL32 gene.

A positive regulator of the ribosomal protein gene, beta factor, belongs to the ETS oncoprotein family.

The beta factor, which interacts with the rpL32 promoter, binds to the sequence 5'-GAGCCGGAAGTG and trans-activates this gene. Comparison of the DNA sequences bound by the beta factor with those bound by other known DNA-binding proteins revealed that the ETS proteins interact with similar DNA sequences. Consequently we have examined the relationship of the beta factor to the several ETS proteins so far reported. Antibody and oligonucleotide competition experiments, performed by using electrophoretic shift analysis, revealed that the beta factor contains ETS epitopes and that it is immunologically related to both of the GA-binding proteins (GABPs), implying that the beta factor may consist of two separate protein subunits.

Characterization of the rRNA-encoding genes and transcripts, and a group-I self-splicing intron in Pneumocystis carinii.

Although Pneumocystis carinii is the most common opportunistic pathogen infecting individuals with AIDS, very little is known of the basic biology of the organism. We have examined the ribosomal RNA (rRNA) and the DNA encoding it (rDNA) in P. carinii in an attempt to clarify its taxonomic position and to begin to study its genetic processes. Electrophoretic analysis showed that the sizes of the P. carinii rRNAs are quite similar to the sizes of the corresponding rRNAs from Saccharomyces cerevisiae. Direct sequence analysis of approx. 60% of the 18S small subunit-rRNA (Ss-rRNA) confirmed that its sequence is similar to that of yeast-like fungi and that a putative group-I intron previously observed in the 18S rDNA is, in fact, excised from the mature rRNA. PCR analysis of the intron in P. carinii genomic DNA showed that each of the multiple rDNA genes bears the group-I intron and in vitro transcripts of the intron autocatalytically excise from the rRNA primary transcript in the presence of GTP. Finally, analogues of GTP inhibit the self-splicing reaction, indicating that the guanosine-binding site of the intron closely resembles that of other well-characterized group-I introns. Since no group-I introns have been found in higher eukaryotes, this self-splicing process represents a viable target for chemotherapy of P. carinii pneumonia (PCP).

A downstream sequence of the rpL32 promoter competes with the glucocorticoid responsive element for a protein factor.

The murine ribosomal protein (rp) L32 gene contains essential promoter sequences located both upstream and downstream of the cap site. A combination of gel mobility shift, UV cross-linking, and cell-free transcription assays were used to analyze the interaction of factors binding to a downstream element (located at position +25 to +37). The rpL32 downstream element identified polypeptides (transcription factors) ranging in size from 45 to 25 kilodaltons (kDa). Four base pair changes in the wild-type sequence of the downstream element eliminated binding. An oligonucleotide containing the glucocorticoid responsive element sequence competed specifically for the 45-kDa protein in both the gel mobility shift assay and in the UV cross-linking studies. Our data also indicate that the downstream binding factors contribute to cell-free transcription of the rpL32 gene.

Yeast transcription factor IID participates in cell-free transcription of a mammalian ribosomal protein TATA-less promoter.

We analysed transcription of the gene for the ribosomal protein (rp) L32 of the mouse, which is transcribed in mouse L1210 nuclear extracts in vitro. The rpL32 gene lacks a canonical TATA box. Hence it has been suggested that this gene has an alternative transcription pathway not requiring transcription factor IID (TFIID). Selective inactivation of TFIID in nuclear extract completely abolished the transcription of rpL32 in vitro. Selective inactivation was restored by the addition of cloned and purified yeast TFIID (yTFIID), indicating that this TATA-less rpL32 promoter utilizes TFIID for its transcription initiation. Furthermore, addition of an oligonucleotide-containing TATA sequence interfered with the rpL32 transcription and this was overcome by the addition of yTFIID. To further examine the stage of involvement of TFIID in rpL32 transcription, TATA oligonucleotide was added to nuclear extract before and after the formation of the transcription complex. The results reveal that TFIID associates with the pre-initiation complex and that this complex is largely resistant to added TATA oligonucleotide. Our results show, for the first time, that the TATA-less rpL32 gene utilizes TFIID for transcription initiation.

Enhanced cell-free transcription of the ribosomal protein L32 gene by the polyoma virus enhancer PEA3 DNA-binding protein.

The mouse-ribosomal-protein-L32-gene promoter contains a 12-bp sequence motif within the 5'-upstream region termed the beta element which shows significant similarity with the consensus sequence of the polyoma-virus-enhancer PEA3. A cloned PEA3 DNA-binding protein, expressed in Escherichia coli and purified, activates the expression of the ribosomal-protein-L32 gene in a cell-free system. Moreover, the PEA3 protein participates in the formation of the ribosomal-protein-L32-promoter-preinitiation-transcription complex. The preinitiation complex formed with PEA3 is resistant to competition by oligonucleotides containing the beta element. In addition anti-PEA3 serum interacts with a factor in mouse L1210 nuclear extract that binds to the beta element, causing a supershift in a mobility-shift assay. Our study demonstrates for the first time that the PEA3 protein can transactivate a cellular gene in a cell-free transcription system.

Identification of a polypeptide bound to the beta region of the mouse r protein L32 promoter.

Studies have been initiated to identify the protein component(s) which interact with the beta regulatory region of the mouse ribosomal protein L32 gene promoter. By the combined use of the mobility shift assay and UV cross-linking, a factor specific for the upstream transcriptional control sequence of the beta region of the ribosomal protein L32 promoter has been detected in mouse L1210 nuclear extracts. A mutation (GT----TC at -71 to -70) in this sequence eliminates the binding. Beta factor is identified as a 55 kDa polypeptide by UV cross-linking. Addition of excess beta element (double-stranded oligonucleotide) to a cell-free transcription system reduces transcription of the ribosomal protein L32 gene. Our results provide evidence that the interaction between the beta element and the beta factor is involved in ribosomal protein L32 transcription.

Histamine blockade and cardiovascular changes following heparin administration during cardiac surgery.

Large doses of heparin given as a bolus may produce hypotension; however, conflicting reports exist about the mechanisms involved. This study was undertaken to determine the role of histamine in beef lung heparin-induced hypotension and the efficacy of histamine-receptor blockade in attenuating this undesirable side effect in patients undergoing cardiac surgery. Two hundred patients with good ventricular function were studied after they were randomized into four equal groups. Group I (control) received no histamine-receptor blockade, group II received 1 mg/kg of diphenhydramine 30 minutes prior to heparin administration, group III received 5 mg/kg of cimetidine 4 hours and again 30 minutes before heparin administration, and group IV received 1 mg/kg of diphenhydramine 30 minutes prior to heparin administration and 5 mg/kg of cimetidine 4 hours and 30 minutes before heparin administration. Hemodynamic variables, plasma histamine, and ionized calcium levels were measured before and after heparin administration. Significant hypotension occurred in group I patients after heparin administration. Mean arterial pressure decreased from 95 +/- 5 to 67 +/- 1.5 mm Hg (P less than 0.005) after 1 minute and to 85 +/- 2 mm Hg (P less than 0.05) at 4 minutes. Those changes were significantly greater than in group II (P less than 0.025) and Group IV (P less than 0.005) patients, in whom no significant hypotension was found. In group III, mean arterial pressure decreased from 92 +/- 3 to 75 +/- 1 mm Hg (P less than 0.05) after 1 minute and returned toward baseline values after 4 minutes. Histamine levels increased significantly in all groups of patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Phlebotomy via the pulmonary artery catheter introducer for intraoperative autotransfusion.

Fear of the acquired immune deficiency syndrome and other blood-transmitted diseases has created a revival of autologous transfusion during cardiac surgery. The present report is of 200 patients undergoing cardiopulmonary bypass during cardiac surgery in whom phlebotomy was performed via the sideport of the introducer for the pulmonary artery catheter for later reinfusion. Each unit of phlebotomized blood was replaced with 500 mL of normal saline. Cardiac output and mean arterial blood pressure decreased significantly after phlebotomy (P less than 0.05) and returned toward control values after administration of the sodium chloride. The autologous blood was replaced after cardiopulmonary bypass. Fresh frozen plasma and platelets were not administered to the patients in the operating room. Eleven patients undergoing coronary artery bypass grafting received fresh frozen plasma in the recovery room because they were receiving aspirin and dipyridamole up to the day of surgery. Prolonged duration of cardiopulmonary bypass in two double-valve replacements, and one coronary artery bypass graft patient who required insertion of an intra-aortic balloon, accounted for the administration of fresh frozen plasma and platelets in three patients. The average volume of phlebotomized blood was 875 mL, which resulted in a decrease of the hematocrit from 40.5% +/- 0.5% (P less than 0.05) to 29.75% +/- 0.5% and 30.5% +/- 0.5% at the end of surgery and at discharge from the hospital, respectively. Phlebotomy via the Y port of the introducer of the pulmonary artery catheter is an easy, simple, and cost-effective way to remove autologous blood in patients undergoing cardiac surgery.

An electrophoretic karyotype and assignment of ribosomal genes to resolved chromosomes of Pneumocystis carinii.

Pulsed-field gel electrophoresis was used to generate a molecular karyotype of chromosomes from the opportunistic AIDS pathogen, Pneumocystis carinii. P. carinii cysts and trophozoites were isolated from immunosuppressed rats, lysed in situ in agarose blocks, and subjected to orthogonal-field gel electrophoresis (OFAGE) and contour-clamped homogeneous-field gel electrophoresis (CHEF). OFAGE and CHEF gels resolved, respectively, 16 or 20 chromosome bands ranging in size from 0.32-1.5 megabase pairs. Summation of the estimated sizes of these chromosomes suggested a total genome complexity for P. carinii of 8-16 megabase pairs. Homologous probes for the genes encoding the 18S, 5.8S, and 5S ribosomal RNAs were hybridized to filter blots of the pulsed-field gels to map these genes to specific P. carinii chromosomes.

Cellular localization and age dependent changes in mRNA for glutathione S-transferase-P in rat testicular cells.

Using Northern blotting techniques we report that mRNA for Glutathione S-transferase-P (GST-P or GST 7-7) is present in rat testis. GST-P mRNA was detected in cultured Sertoli cells, cultured peritubular cells, as well as in transplantable Leydig cell tumor. However, no GST-P mRNA was detected in rat germ cell fractions. There was a marked increase in mRNA for GST-P from day 5 to day 20 in rats, after which a decrease was seen. The decreased level of mRNA for GST-P in the testis after 20 days of age, coincided in time with the exponential increase in germ cells, and accompanying relative decrease in somatic cells. The results show that mRNA for GST-P is primarily present in somatic cells of the rat testis.

Hemodynamic changes after nafcillin administration during coronary artery bypass surgery.

The hemodynamic response to nafcillin administration was studied in 45 patients with good left ventricular function and no known history of hypersensitivity to penicillin during coronary artery bypass grafting (CABG). Group I (15 patients) received 1 gram of nafcillin in 10 mL of saline as an intravenous (IV) bolus, group II (15 patients) received 1 gram of nafcillin in 50 mL of saline as a slow IV infusion over 15 minutes, and group III (15 patients) did not receive nafcillin. Hemodynamic variables and plasma histamine and catecholamine levels were measured before and after nafcillin administration, after 500 mg of CaCl2, and after 0.1 mg of phenylephrine. Bolus nafcillin administration produced profound hypotension secondary to vasodilatation with significant increases in cardiac index and decreases in systemic and pulmonary vascular resistances. Cardiac index increased from 3.15 +/- 0.3 L/min/m2 to 5.75 +/- 0.25 L/min/m2 (P less than 0.005) one minute after nafcillin administration, and remained at 5.1 +/- 0.35 L/min/m2 after administration of CaCl2 (P less than 0.005). All hemodynamic parameters returned toward control values after administration of 0.1 mg of phenylephrine, IV. Plasma epinephrine, norepinephrine, and histamine levels increased more than 100%. In group II, cardiac index increased, while systemic and pulmonary vascular resistances and mean arterial pressure decreased. However, these changes were less significant than those found in group I.(ABSTRACT TRUNCATED AT 250 WORDS)

Investigation of detoxification capacity of rat testicular germ cells and Sertoli cells.

Isolated pachytene spermatocytes liver longer than round spermatids in vitro. Indigenous formation of oxygen-derived free radicals and hydrogen peroxide can cause damage to germ cells. The germ cell antioxidant capacity may play an important role in this respect. In view of this, we have examined the activity and cellular localization of superoxide dismutase (SOD) and glutathione S-transferases (GST) in rat testicular cells. We have found significant differences in the distribution of these enzymatic activities in the germ cells. In addition, this study shows that alpha-tocopherol is found in various amounts in rat testicular cells in the order of: Sertoli cells greater than pachytene spermatocytes greater than round spermatids, with a factor of 4 in the alpha-tocopherol content between Sertoli cells and round spermatids.

Comparison of superoxide dismutase, thiopental, and nimodipine for maintenance of somatosensory evoked responses during aortic cross-clamping and declamping in dogs.

Paraplegia is a potential complication of aortic cross-clamping. The occurrence of this devastating sequela has caused increased interest in the use of somatosensory evoked responses (SER) to monitor spinal cord ischemia during aortic cross-clamping. This study was designed to examine changes in SERs during clamping and declamping of the canine aorta after injection of superoxide dismutase (SOD), thiopental (T), and nimodipine (N). In the control group, cross-clamping the aorta produced an increase in latency and a decrease in amplitude of the SER starting at two minutes. Isoelectric SERs were obtained after 16 minutes of aortic cross-clamping, but recovered with cross-clamp removal. When the aorta was clamped for more than 16 minutes in the control group, the isoelectric SERs obtained were irreversible. After the injection of SOD and T, SER latencies and amplitudes changed to a smaller degree with aortic cross-clamping and did not become isoelectric even after 20 minutes of clamping. During aortic cross-clamp removal in the control group, SERs initially improved and then showed signs of reperfusion ischemia, which disappeared after eight minutes. There were no significant SER changes due to reperfusion when SOD or T or the combination was given prior to aortic cross-clamping. There was no difference in SER changes from the control group during aortic cross-clamping and after release of cross-clamping when N was given. Nimodipine did not alter SER changes from aortic cross-clamping alone. In summary, SOD and T, alone or in combination, protect the spinal cord against ischemia during aortic cross-clamping and declamping.