Diffusion- and perfusion-weighted brain magnetic resonance imaging in patients with neurologic complications after cardiac surgery.

BACKGROUND: Neurologic complications after cardiac surgery include stroke, encephalopathy, and persistent cognitive impairments. More precise neuroimaging of patients with these complications may lead to a better understanding of the etiology and treatment of these disorders. OBJECTIVE: To study the pattern of ischemic changes on diffusion- and perfusion-weighted magnetic resonance imaging (DWI, and MRPI, respectively) in patients with neurologic complications after cardiac surgery. METHODS: All records were reviewed of our patients undergoing cardiac surgery in the previous year who also underwent postoperative DWI or MRPI. Neurologic symptoms, vascular studies, and the pattern of ischemic changes were recorded. Acute ischemic lesions were classified as having a territorial, watershed, or lacunar pattern of infarction. Patients with multiple territorial infarcts in differing vascular distributions that were not explained by occlusive vascular lesions were classified as having multiple emboli. RESULTS: Fourteen patients underwent DWI and 4 underwent MRPI. Acute infarcts were found in 10 of 14 patients by DWI as compared with 5 of 12 patients by computed tomography. Eight patients presented with encephalopathy (associated with focal neurologic deficits in 4), 4 with focal deficits alone, and 2 with either fluctuating symptoms or transient ischemic attacks. Among patients with encephalopathy, 7 of 8 had patterns of infarction suggestive of multiple emboli, including 3 of 4 patients with no focal neurologic deficits. Several patients had combined watershed and multiple embolic patterns of ischemia. Findings of MRPI studies were abnormal in 2 of 4 patients, showing diffusion-perfusion mismatch; both patients had either fluctuating deficits or transient ischemic attacks, and their conditions improved with blood pressure manipulation. CONCLUSIONS: In patients with neurologic symptoms after cardiac surgery, DWI is more sensitive to ischemic change than computed tomographic scanning and can demonstrate patterns of infarction that may help us understand etiology. The most common pattern was multiple embolic infarcts. Preliminary experience with MRPI suggests that some patients have persistent diffusion-perfusion mismatch after surgery and may benefit from therapeutic intervention.

Paper-based archiving of mammalian and plant samples for RNA analysis.

The ability to archive biological samples for subsequent nucleic acid analysis is essential for tissue specimens and forensic samples. FTA Card is a chemically treated filter paper designed for the collection and room temperature storage of biological samples for subsequent DNA analysis. Its usefulness for the preservation of biological samples for subsequent RNA analysis was tested. Here, we demonstrate that RNA in biological samples stored on FTA Cards is stable and can be used successfully for RT-PCR and northern blot analysis. RNA stability depends on the storage temperature and the type of biological specimen. RNA in mammalian cells stored on FTA Cards is stable for over one year at temperatures at or below -20 degrees C and for two to three months in samples stored at room temperature. For plant leaf, longer storage times (> 5 days) require temperatures at or below -70 degrees C following sample application. FTA Cards may constitute a method not only for convenient collection and storage of biological samples but also for rapid RT-PCR analysis of tissue and cell samples.

Long PCR for VNTR analysis.

The Polymerase Chain Reaction (PCR) has revolutionized the analysis of DNA from a variety of sources. With its sensitivity and ability to amplify degraded DNAs and small quantities of samples, coupled with fast turn-around-time, PCR is often the analytical method of choice for DNA profiling in forensic laboratories. RFLP methods, while requiring larger amounts of high molecular weight DNA and needing approximately 6-8 weeks of analytical time, still provide a higher power of discrimination per locus than that achieved using the loci currently available for PCR. The combination of both RFLP and PCR would be advantageous for some applications. A new technique, Long PCR, allows for the effective amplification of long DNA targets from approximately 0.5 kb to > 20 kb of genomic DNA. Currently, several Long PCR systems are commercially available. Using a Taq/Pyrococcus DNA polymerase enzyme system and DNA isolated from bloodstains, we have successfully amplified 1-20 ng of Chelex-extracted DNA, an amount commonly used in Amp-FLP technology. The robustness of Long PCR in comparison to RFLP was also examined through the use of partially degraded blood samples. Long PCR was then used to amplify both D2S44 and D5S110 RFLP loci. Although all D2 and D5 alleles were detected, the larger alleles were amplified at significantly lower levels than the smaller alleles.

Determinants of cognitive change after coronary artery bypass surgery: a multifactorial problem.

BACKGROUND: Several studies have investigated predictors of cognitive decline after coronary artery bypass grafting (CABG), but there is little consensus as to which specific factors are predictive of poor cognitive outcomes. METHODS: We evaluated 127 patients undergoing CABG with standardized neuropsychological tests preoperatively, at 1 month and at 1 year. The outcome measure was a continuous variable reflecting change in z-scores for eight cognitive domains over time for individual patients. Univariate analyses were performed to evaluate the association between the demographic, operative, and postoperative factors and the cognitive outcome variables. Factors that were significant were included in a multiple linear regression analysis. RESULTS: Among the medical history variables, diabetes was associated with change in executive functions and psychomotor speed. Some of the operative variables were associated with short-term changes, but none with the 1-year outcomes. For example, the surgeon's rating of degree of difficulty in selecting a cross-clamp site was associated with change in attention. Higher mean pump rate during the procedure was associated with improved performance on tests of language. The cognitive domains associated with medical variables were different from those associated with surgical variables, and the associations observed at 1-year were different from those seen at 1-month. CONCLUSIONS: Change in cognition after CABG is associated with both medical and surgical variables. The specifics of these associations depend on the choice of time points after surgery. This suggests that there are multiple etiologies for these changes, with nonspecific effects of anesthesia and prolonged surgery interacting with the more specific effects of the surgical procedure itself.

Assessing the impact of cerebral injury after cardiac surgery: will determining the mechanism reduce this injury?

BACKGROUND: Central nervous system dysfunction continues to produce significant morbidity and associated mortality in patients undergoing cardiac surgery. Using a closed-chest canine cardiopulmonary bypass model, dogs underwent 2 h of hypothermic circulatory arrest (HCA) at 18 degrees C, followed by resuscitation and recovery for 3 days. Animals were assessed functionally by a species-specific behavioral scale, histologically for patterns of selective neuronal necrosis, biochemically by analysis of microdialysis effluent, and by receptor autoradiography for N-methyl-D-aspartate (NMDA) glutamate receptor subtype expression. RESULTS: Using a selective NMDA (glutamate) receptor antagonist (MK801) and an AMPA antagonist (NBQX), glutamate excitotoxicity in the development of HCA-induced brain injury was documented and validated. A microdialysis technique was employed to evaluate the role of nitric oxide (NO) in neuronal cell death. Arginine plus oxygen is converted to NO plus citrulline (CIT) by the action of NO synthase (nNOS). CIT recovery in the cerebrospinal fluid and from canine cortical homogenates increased during HCA and reperfusion. These studies demonstrated that neurotoxicity after HCA involves a significant and early induction of nNOS expression, and neuronal processes leading to widespread augmentation of NO production in the brain. To further investigate the production of excitatory amino acids in the brain, we hypothesized the following scenario: HCA--> increased glutamate, increased aspartate, increased glycine--> increased intracellular Ca2+--> increased NO + CIT. Using the same animal preparation, we demonstrated that HCA caused increased intracerebral glutamate and aspartate that persists up to 20 h post-HCA. HCA also resulted in CIT (NO) production, causing a continued and delayed neurologic injury. Confirmatory evidence of the role of NO was demonstrated by a further experiment using a specific nNOS inhibitor, 7-nitroindazole. Animals underwent 2 h of HCA, and then were evaluated both physiologically and for NO production. 7-Nitroindazole reduced CIT (NO) production by 58.4 +/- 28.3%. In addition, dogs treated with this drug had superior neurologic function compared with untreated HCA controls. CONCLUSIONS: These experiments have documented the role of glutamate excitotoxicity in neurologic injury and have implicated NO as a significant neurotoxin causing necrosis and apoptosis. Continued research into the pathophysiologic mechanisms involved in cerebral injury will eventually yield a safe and reliable neuroprotectant strategy. Specific interventional agents will include glutamate receptor antagonists and specific neuronal NO synthase inhibitors.

Neurobehavioural sequelae of cardiopulmonary bypass.

The development of coronary artery bypass grafting (CABG) and its effect on angina is the product of a series of technical and scientific advances. Despite these advances, however, adverse neurobehavioural outcomes continue to occur. Stroke is the most serious complication of CABG, but studies that have identified demographic and medical risk factors available before surgery are an important advance. Short-term cognitive deficits are common after CABG, but may not be specific to this procedure. However, deficits in some cognitive areas such as visuoconstruction persist over time, and may reflect parieto-occipital watershed area injury secondary to hypoperfusion or embolic factors. Risk factors for cognitive decline may be time dependent, with short-term studies identifying factors that differ from those of long-term studies. Patients with depression before surgery are likely to have persistent depression afterwards. However, depression does not account for the cognitive decline after CABG. Since CABG is increasingly done in older patients with more comorbidity, the challenge is to identify patients at risk of adverse neurocognitive outcomes and to protect them by modification of the surgical procedure or by effective medical therapy.

Prevalence of leg wound complications after coronary artery bypass grafting: determination of risk factors.

BACKGROUND: The reported prevalence of leg wound complications after coronary artery bypass grafting is 2% to 24%. Decreased length of hospital stay for patients who have this surgical procedure poses new care requirements in both acute care and community settings. OBJECTIVE: To determine the prevalence of postoperative leg wound complications in patients undergoing coronary artery bypass grafting and the risk factors associated with these complications. METHOD: In this prospective, observational study, 547 consecutive patients who had coronary artery bypass grafting alone or in combination with other cardiac surgical procedures were examined for evidence of leg wound complications each day after surgery during hospitalization. After discharge, problems were detected by home care nurses. RESULTS: The prevalence of leg wound complications was 6.8%. Factors significant by multiple logistic regression included preoperative hospitalization, use of an Ace elastic bandage in the operating room, the length of time the leg incision remained open in the operating room, and administration of nicardipine intravenously in the intensive care unit. Odds ratios were calculated for each variable. Premorbid factors such as diabetes or peripheral vascular disease were not predictive of complications. On average, most problems occurred on postoperative day 10, when many patients were at home. CONCLUSIONS: The results highlight the need to detect complications early, in both the hospital and the community settings. The determination of factors related to poor outcomes may assist clinicians in improving healthcare delivery.

Depression and cognitive decline after coronary artery bypass grafting.

BACKGROUND: Depression is commonly reported after coronary artery bypass grafting (CABG), and after cardiac surgery in general. Many earlier reports relied on non-standard assessments of depression, which may have overestimated its frequency. Cognitive decline has also been reported after CABG. We assessed the frequency of depression after CABG by a validated depression measure (Center for Epidemiological Study of Depression, CES-D), and examined the relation between depression and cognitive decline. METHODS: Patients were tested before CABG and 1 month and 1 year after surgery with a series of neuropsychological tests that assessed a range of cognitive areas. Depressed mood was measured by the CES-D scale, and defined as a score above 16. FINDINGS: 90 (73%) of the 124 patients were not depressed before surgery, and 34 were depressed at that time. Only 12 (13%) of patients not depressed before surgery were depressed at 1 month afterwards, whereas 18 (53%) of those who were depressed before surgery were depressed at 1 month (p < 0.001). 8 (9%) patients not depressed before surgery were depressed at 1 year; 16 (47%) of patients who were depressed before CABG were depressed at 1 year (p < 0.001). Statistical analysis showed only minimal correlation-or none at all-between depression and eight areas of cognitive outcome, or between changes in depressed status and cognitive scores. INTERPRETATION: Of those patients who were depressed after CABG, the large majority were depressed before surgery. There was no correlation, moreover, between depressed mood and cognitive decline after CABG, which suggests that depression alone cannot account for cognitive decline.

Cognitive outcome after coronary artery bypass: a one-year prospective study.

BACKGROUND: Cognitive deficits have been reported in patients after coronary artery bypass grafting, but the incidence of these deficits varies widely. We studied prospectively the incidence of cognitive change and whether the changes persisted over time. METHODS: Cognitive testing was done preoperatively and 1 month and 1 year postoperatively in 127 patients undergoing coronary artery bypass grafting. Tests were grouped into eight cognitive domains. A change of 0.5 standard deviation or more at 1 month and 1 year from patient's preoperative Z score was the outcome measure. RESULTS: We identified four main outcomes for each cognitive domain: no decline; decline and improvement; persistent decline; and late decline. Only 12% of patients showed no decline across all domains tested; 82% to 90% of patients had no decline in visual memory, psychomotor speed, motor speed, and executive function; 21% and 26% had decline and improvement in verbal memory and language; approximately 10% had persistent decline in the domains of verbal memory, visual memory, attention, and visuoconstruction; and 24% had late decline (between 1 month and 1 year) in visuoconstruction. CONCLUSIONS: This study establishes that the incidence of cognitive decline varies according to the cognitive domain studied and that some patients have persistent and late cognitive changes in specific domains after coronary artery bypass grafting.

Predictors of stroke risk in coronary artery bypass patients.

BACKGROUND: Stroke occurs after coronary artery bypass grafting with an incidence ranging between 0.8% and 5.2%. To identify factors associated with stroke, we prospectively examined a study cohort and tested findings in an independent validation sample. METHODS: The study cohort comprised 456 patients undergoing coronary artery bypass grafting only, and the validation sample comprised 1,298 patients. Stroke was detected postoperatively by the study team and confirmed by neurologic consultation and computed tomographic scanning. RESULTS: Five factors taken together were correlated with stroke: previous stroke, presence of carotid bruit, history of hypertension, increasing age, and history of diabetes mellitus. The only significant intraoperative factor was cardiopulmonary bypass time. Probabilities were calculated, and patients were placed into low, medium, and high stroke-risk groups. In the validation sample, this model was able to rank the majority of patients with stroke into the high-risk group. CONCLUSIONS: These five factors taken together can identify the risk of stroke in patients having coronary artery bypass grafting. Recognition of the high-risk group will aid studies on the mechanism and prevention of stroke by modification of surgical procedures or pharmacologic intervention.

Neurologic injury in cardiac surgical patients with a history of stroke.

BACKGROUND: Controversy still exists as to whether patients with previous stroke are at increased risk for neurologic complications after heart operations. METHODS: We performed a prospective analysis of 1,000 consecutive patients undergoing cardiac operations requiring cardiopulmonary bypass, without hypothermic circulatory arrest. Of the 1,000 patients, 71 had previously documented stroke (study group); 2 control patients with no history of stroke were selected for each of these patients (control group, n = 142). There were no significant differences between the study and control patients with respect to established risk factors for neurologic complications. RESULTS: Compared with controls, study patients took longer to awaken (12.6 +/- 10.9 versus 3.5 +/- 2.1 hours; p < 0.0001) and longer to extubate (29.5 +/- 29.3 versus 9.1 +/- 5.2 hours; p < 0.001), and had a greater incidence of reintubation (7 of 71, 9.9% versus 2 of 142, 1.4%; p < 0.01) and postoperative confusion (26 of 71, 36.6% versus 7 of 142, 4.9%; p < 0.001). There was a higher incidence of focal neurologic deficit among study patients (31 of 71, 43.7% versus 2 of 142, 1.4%; p < 0.001). These deficits included new stroke (6 of 71, 8.5%) as well as the reappearance of previous deficits (19 of 71, 26.8%) or worsening of previous deficits (6 of 71, 8.5%), without new abnormalities on head computed tomography or magnetic resonance imaging. Study patients with neurologic deficit had longer cardiopulmonary bypass times than did study patients without deficit (146 +/- 48.5 versus 110 +/- 43.3 minutes; p < 0.001). The 30-day mortality rate was greater in study patients than in controls (5 of 71, 7% versus 1 of 142, 0.7%; p < 0.02), with four deaths among the 6 study patients with a new stroke (66.7%). CONCLUSION: This analysis identifies a group of patients at high risk for neurologic sequelae and confirms the vulnerability of the previously injured brain to cardiopulmonary bypass, as evidenced by reappearance or exacerbation of focal deficits in such patients.

Neuropsychologic change after cardiac surgery: a critical review.

Studies that have examined neuropsychologic change after cardiac surgery address three main issues: (1) the incidence of cognitive change; (2) the identification of factors that put patients at higher risk; and (3) the evaluation of interventions to prevent these complications. This review attempts to bring together concerns associated with various study designs and to integrate the conclusions from these studies. Thirty-five studies have been examined in this review. Some of the difficulties encountered when quantifying the degree of cognitive change are related to study design, patient sampling, and deficit definition. Additionally, changing patient populations have influenced results reported from different health care settings. Increasing age and longer cardiopulmonary bypass times have been correlated with cognitive decline in a number of studies. Filtration devices and blood gas management techniques have decreased but not eliminated the number of patients who have cognitive decline. Cognitive change exists following cardiac procedures. Identification of a subgroup of patients at high risk for cognitive change has been difficult, possibly due to issues of study design. Design of future studies, which may include intraoperative or pharmacologic interventions, is dependent on identification of this high-risk group.

A comparison study of human papillomavirus prevalence by the polymerase chain reaction in low risk women and in a gynaecology referral group at elevated risk for cervical cancer.

The reported prevalence of human papillomavirus (HPV) type 16 in the genital tracts of women with various gynaecological conditions is highly variable. In particular, some results with the polymerase chain reaction (PCR) technique have suggested that HPV-16 is a ubiquitous or very common virus. We undertook this study to help clarify the current confusion. PCR with HPV consensus L1 primers and specific E6 primers was used to study 89 women attending two gynaecology referral clinics, as well as 99 women attending a health maintenance organization (HMO) clinic; 70 of these latter women had no current or prior history of genital HPV disease. HPV-16 was detected in less than 5% of cytologically normal women from either group and in 17% (6/36) and 31% (9/29) of women with cervical intraepithelial neoplasia (CIN) from the referral clinic and the HMO, respectively. The other high-risk or intermediate-risk HPVs (types 18, 31, 33 or 35) were less prevalent than HPV 16 in all groups of women. A majority of the HPV types detected by the L1 primers in normal women were uncharacterized HPVs. Overall these uncharacterized HPVs were detected in 37% (46/123) of the normal women and in 48% (31/65) of the women with CIN. Using the most sensitive PCR product detection method employed in the study, HPV DNA was detected in 36% (4/11) of swab specimens obtained from the external abdomen.

Human papillomavirus type 56: a new virus detected in cervical cancers.

A new human papillomavirus type (HPV-56) was identified by low stringency Southern blot analysis with an HPV-31 DNA probe, in a cervical intraepithelial neoplasm (CIN). The DNA of this virus was molecularly cloned and shown to be a new HPV type based on the absence of cross-reactivity to HPV types 1 to 55 under high-stringency hybridization conditions. At low stringency, HPV-56 was most related to HPV types 30 and 45. The deduced organization of the open reading frames of HPV-56, from hybridization and partial nucleotide sequence analyses, reveals a typical HPV genome. HPV-56 was detected in two of 464 normal cervical tissues, in five of 227 cervical condylomas and CIN, and in two of 84 invasive cancers of the cervix.

Nucleotide sequence of human papillomavirus type 31: a cervical neoplasia-associated virus.

The nucleotide sequence of human papillomavirus (HPV) 31 DNA (7912 bp) was determined and used to deduce the genomic organization of this cervical cancer-associated virus. Based on comparisons of the HPV 31 DNA sequence to other sequenced HPVs, HPV 31 is a typical papillomavirus most related to HPV 16 (70% identical nucleotides). The E6 and E7 open reading frames (ORF) of HPV 31 contain several potential DNA binding motifs (Cys-X-X-Cys), the locations of which are conserved in all HPVs. The E6 ORF also has the potential to code for an E6* protein. The E7 ORF of HPV31 encodes a polypeptide motif which appears to distinguish HPVs associated with cervical cancer, such as types 16, 18, 31, and 33, from HPVs found primarily in benign lesions, such as types 6 and 11.

Cloning and partial DNA sequencing of two new human papillomavirus types associated with condylomas and low-grade cervical neoplasia.

Using low-stringency Southern blot analysis and cloning in lambda bacteriophage, two new human papillomavirus types (HPV-43 and HPV-44) were identified and their DNAs were cloned from vulvar tissues. The isolates were characterized by restriction endonuclease mapping and shown to be new HPV types on the basis of their minimal hybridization with all other known HPV types at high stringency. Both HPVs are most closely related to types 6, 11, and 13. HPV-43 did not exhibit any cross-reactivity with these HPV types at high stringency. HPV-44 showed minimal cross-reactivity to HPV-13, which was in the range of 20 to 25% according to liquid hybridization analysis. The deduced genomic organization of each of the two new HPVs was colinear with HPV-6b. Prevalence studies revealed that HPV-43 and HPV-44 together were found in 6 of 439 normal cervical tissues, in 8 of 195 cervical intraepithelial neoplasms, but in none of 56 cervical cancers tested thus far.

Characterization of murine A-raf, a new oncogene related to the v-raf oncogene.

A 1.6-kilobase cDNA (A-raf) has been isolated from a murine spleen cDNA library which encodes part of a protein related to the raf oncogene. Its amino acid sequence has 85% homology to raf in a central portion of 100 amino acids. In contrast to raf, A-raf shows a highly restricted tissue distribution of expression, with highest levels observed in epididymis, followed by intestine. When incorporated into a retrovirus, the resulting gag-A-raf fusion gene causes transformation in vitro and induces tumors in newborn mice. Thus, A-raf represents a new proto-oncogene. Transformation by A-raf is independent of ras gene function, as is the case for raf and mos but not other oncogenes.

Actively transcribed genes in the raf oncogene group, located on the X chromosome in mouse and human.

Murine and human cDNAs, related to but distinct from c-raf-1, have been isolated and designated mA-raf and hA-raf, respectively. The mA-raf and hA-raf cDNAs detect the same murine and human fragments in Southern blots of restriction enzyme-cleaved murine and human cellular DNA. The murine restriction enzyme fragments homologous to mA-raf cDNA cosegregate with mouse chromosome X in a panel of Chinese hamster-mouse hybrid cells, thus localizing the mA-raf locus to mouse chromosome X. Two independently segregating loci, detected by the hA-raf cDNA (or mA-raf cDNA), hA-raf-1 and hA-raf-2, are located on human chromosomes X and 7, respectively. The mA-raf locus and the hA-raf-1 locus are actively transcribed in several mouse and human cell lines.

Structure and biological activity of v-raf, a unique oncogene transduced by a retrovirus.

We have molecularly cloned a unique acutely transforming replication-defective mouse type C virus (3611-MSV) and characterized its acquired oncogene. The viral genome closely resembles Moloney (M) murine leukemia virus (MuLV), except for a substitution in M-MuLV in the middle of p30 and the middle of the polymerase gene (pol). Heteroduplex analysis revealed that 2.4 kilobases of M-MuLV DNA were replaced by 1.2 kilobases of cellular DNA. The junctions between viral and cellular sequences were determined by DNA sequence analysis to be 517 nucleotides into the p30 sequence and 1,920 nucleotides into the polymerase sequence. Comparison of the transforming gene from 3611-MSV, designated v-raf, with previously isolated retrovirus oncogenes either by direct hybridization or by comparison of restriction fragments of their cellular homologs shows it to be unique. Transfection of NIH 3T3 cells with cloned 3611-MSV proviral DNA leads to highly efficient transformation and the recovered virus elicits tumors in mice typical of the 3611-MSV virus. Transfected NIH 3T3 cells express two 3611-MSV-specific polyproteins (P75 and P90), both of which contain NH2-terminal gag gene-encoded components linked to the acquired sequence (v-raf) translational product. The cellular homolog, c-raf, is present in one or two copies per haploid genome in mouse and human DNA.