Immunohistochemical assessment of rat nerve isografts and immunosuppressed allografts

Objective: Autologous peripheral nerve grafts are commonly used clinically as a treatment for peripheral nerve injuries. However, in research using an autologous graft is not always feasible due to loss of function, which in many cases is assessed to determine the efficacy of the peripheral nerve graft. In addition, using allografts for research require the use of an immunosuppressant, which creates unwanted side effects and another variable within the experiment that can affect regeneration. The objective of this study was to analyze graft rejection in peripheral nerve grafts and the effects of cyclosporine A (CSA) on axonal regeneration.

Methods: Peripheral nerve grafts in inbred Lewis rats were compared with Sprague-Dawley (SD) rats to assess graft rejection, CSA side effects, immune responses, and regenerative capability. Macrophages and CD8+ cells were labeled to determine graft rejection, and neurofilaments were labeled to determine axonal regeneration.

Results: SD rats without CSA had significantly more macrophages and CD8+ cells compared to Lewis autografts, Lewis isografts, and SD allografts treated with CSA. Lewis autografts, Lewis isografts, and SD autografts had significantly more regenerated axons than SD rat allografts. Moreover, allografts in immunosuppressed SD rats had significantly less axons than Lewis rat autograft and isografts.

Discussion: Autografts have long been the gold standard for treating major nerve injuries and these data suggest that even though CSA is effective at reducing graft rejection, axon regeneration is still superior in autografts versus immunosuppressed allografts.     

Repression of proinflammatory gene expression by lipid extract of Nostoc commune var sphaeroides Kützing,a blue-green alga,via inhibition of nuclear factor-κB in RAW 264.7 macrophages

We investigated whether lipid extract from a blue-green alga, N commune, modulates proinflammatory gene expression in RAW 264.7 macrophages. The cells were incubated with N commune lipid extract (0-100 μg/mL) and subsequently activated by LPS (100 ng/mL). Quantitative real-time PCR analysis showed that mRNA abundance of proinflammatory mediators, including TNF-α, COX-2, IL-1β, IL-6, and iNOS, was significantly reduced by N commune lipid extract in a dose-dependent manner. Secretion of TNF-α and IL-1β into cell culture medium was also significantly decreased by N commune lipid extract. Thin-layer chromatography–densitometry analysis showed that N commune lipid extract contained approximately 15% of fatty acids. To determine whether the inhibition of proinflammatory mediator production by N commune lipid extract is primarily conferred by fatty acids in the lipid extract, macrophages were incubated with 100 μg/mL of N commune lipid extract or 15 μg/mL of a fatty acid mixture, which was formulated to reflect the fatty acid composition of N commune lipid extract. The fatty acid mixture significantly reduced RNA abundance of TNF-α and COX-2, but to a lesser extent than did the N commune lipid extract, suggesting the presence of additional bioactive compounds with an antiinflammatory property in the lipid extract. As NF-κB is a major regulator for the proinflammatory gene expression, we measured its DNA-binding activity. DNA-binding activity of NF-κB was significantly reduced by N commune lipid extract. In conclusion, our study suggests that N commune lipid extract represses the expression of proinflammatory genes in RAW 264.7 macrophages, at least in part, by inhibiting the activation of NF-κB pathway.     

Cardiac issues in the pediatric emergency room.

A great variety of problems referable to the cardiovascular system can prompt a visit to the pediatric emergency room. From the initial presentation of congenital heart disease, to the subsequent life-long management of these patients, to miscellaneous problems like Kawasaki disease and chest pain, the front-line pediatrician must be skilled in the recognition and early management of myriad complaints. This article focuses on information that can assist the emergency pediatrician in the evaluation and treatment of the cardiac patient from arrival in the emergency room until transfer of care to the pediatric cardiologist or inpatient staff.     

Visual-evoked potentials: assessment of retrobulbar and peribulbar anesthesia

PURPOSE: To assess the effects of retrobulbar and peribulbar anesthesia on nerve function as detected by visual-evoked potentials (VEPs). SETTING: University hospital in southern Brazil. METHODS: In a prospective study, 7 patients had peribulbar anesthesia and 9 had retrobulbar anesthesia for extracapsular cataract extraction. Visual-evoked potentials with pattern reversal and flash stimulation were performed at least 1 month before and 1 month after surgery. Study participants did not have ocular pathology other than cataract. The Lens Classification System III was used to grade the opacities before surgery. RESULTS: No significant difference was found between preoperative and postoperative evaluations in VEP flash and pattern-reversal amplitude and latency in either group (P >.05). Postoperative amplitude and latency was not significantly different between the peribulbar and retrobulbar groups. Two cases in the peribulbar group had altered wave morphology without clinical manifestation postoperatively. All patients had a final best spectacle-corrected visual acuity of 20/20. CONCLUSION: Block anesthetic procedures were safely used in cataract surgery, with no clinical sequelae to the optic nerve.     

Comparison of endothelial cell density estimated by contact and non-contact specular microscopy

PURPOSE: To compare a contact and a non-contact specular microscope in the determination of endothelial cell density. SUBJECTS AND METHODS: One hundred and twenty-one eyes from 70 patients who had undergone various degrees of photorefractive keratectomy for myopia were included. The endothelium was imaged by contact (Konan Clinical Specular Microscope) and non-contact (Topcon SP-1000) specular microscopy and the endothelial cell density estimated. RESULTS: The average endothelial cell density achieved by the contact specular microscope was 3011+/-298 cells/mm2 (mean+/-SD, n=121) and by the non-contact specular microscope 3015+/-265 cells/mm2 (n= 121). The difference in endothelial cell density between the contact and the non-contact specular microscope (contact minus non-contact) was -4+/-175 cells/mm2 (t=0.26, 2p>0.05 in a paired t-test). The sampling error on the estimated endothelial cell density was 76 cells/mm2 for the contact specular microscope and 74 cells/mm2 for the non-contact specular microscope. CONCLUSION: The average endothelial cell density and the precision of the measuring technique were similar for the contact and the non-contact specular microscope. Furthermore, the endothelial cell densities estimated by the two instruments at various values of anterior central corneal refractive power and central corneal thickness were similar. The two instruments can be used interchangeably.     

Prolonged persistence on the ocular surface of fortified gentamicin ointment as compared to fortified gentamicin eye drops

PURPOSE: A comparative study on the elimination of gentamicin from the ocular surface and the concentration of gentamicin in the anterior chamber following application of either an ointment or eye drops containing equal concentrations (1.5%) of gentamicin. METHODS: A disc-diffusion test was used to determine the concentration of gentamicin in fornix inferior of 10 persons. The anterior chamber concentration of gentamicin was determined in 5 cataract patients by the TDX analyzer, Abbot Laboratories, II., USA. RESULTS: Ten minutes following application, the concentration of gentamicin was significantly higher in the eyes receiving ointment (310.6 mg/L) compared to drops (45 mg/L) (p<0.01). Furthermore, gentamicin could be detected 40 minutes after application in the eyes receiving ointment compared to 10 minutes in the eyes receiving drops. The anterior chamber concentration of gentamicin after application of either drops or ointment was lower than 0.6 mg/L and thus below detection limit. CONCLUSIONS: The persistence of gentamicin ointment was significantly longer on the ocular surface as compared to gentamicin eye drops. Gentamycin ointment may thus provide a means to reduce the high application frequency presently in use with eye drops to treat bacterial keratitis and thereby reduce patient inconvenience, especially during nighttime.     

Refractive results of radial keratotomy: a ten-year retrospective study

PURPOSE: To investigate the long-term effects and stability of refraction after radial keratotomy procedure. METHODS: Radial keratotomy was performed on 123 persons to reduce myopia (range: -1 to -13 diopters) in 1986 to 1989. A mean of 11.5 years later (range 10 to 13), 61 of these patients with 102 eyes underwent a standardised refractive examination where subjective spherical equivalent refraction was measured and compared to the preoperative and the one month postoperative refractive measurement collected from the patients records. RESULTS: There was a reduction in spherical equivalent from an average of -5.46 diopters (SD 2.38) preoperatively to -2.32 diopters (SD 1.96) 11.5 years postoperatively. The mean change in direction of myopia between 1 month and 11.5 years postoperatively was 0.17 diopters (SD 1.18). This change was not statistically significant. From 1 month to 11.5 years, 10 of the eyes had developed more than 1 diopter hyperopia, and 20% more than 1 diopter myopia. When asked directly, all patients were satisfied with the result of their operation in general; 2 patients still complained of glare. CONCLUSION: No significant changes in refraction were found between 1 month and 11.5 years after radial keratotomy. Previously reported long-term studies on this field have found a trend toward progressive hyperopia. No evidence of such change can be supported by this study.     

Neuropeptide Y administration into the third ventricle does not increase sucrose or ethanol self-administration but does affect the cortical EEG and increases food intake

RATIONALE: Several studies have provided indirect evidence that neuropeptide Y (NPY) may play a role in the regulation of ethanol consumption. However, the direct effects of central NPY administration on ethanol drinking are unclear. OBJECTIVE: This study examined the effects of NPY on ethanol, sucrose, and food consumption as well as its concomitant effects on the cortical EEG. METHODS: Wistar rats were implanted with cortical recording electrodes and a cannula in the third ventricle after using a sucrose substitution procedure to establish ethanol self-administration. NPY (0-15 microg/3.0 microl) was infused into the third ventricle prior to drinking sessions, when 10% ethanol (10E), 2% sucrose (2S), 0.5% sucrose (0.5S), or food were available. Behavior and cortical EEG were monitored during the sessions. RESULTS: NPY had no effect on the intake of 10E, 2S, or 0.5S, but NPY (15 microg/3.0 microl) significantly increased food intake. Under baseline drinking conditions, EEG power in the 6-8 Hz range was significantly greater when 2S was consumed compared to 10E. NPY decreased power in the 8-16 Hz range, decreased peak frequency in the 6-8 Hz range, and increased peak frequency in the 32-50 Hz range when 10E or 2S was available. CONCLUSIONS: These data suggest that NPY administration into the third ventricle preferentially regulates feeding compared to ethanol or sucrose drinking. In addition, since NPY significantly altered the cortical EEG in the absence of effects on ethanol and sucrose consumption, these data may indicate that NPY's cortical EEG effects are more related to its sedative or anxiolytic properties, rather than any effect on consumption.     

Protective association of genetic variation in alcohol dehydrogenase with alcohol dependence in Native American Mission Indians

OBJECTIVE: Two alcohol dehydrogenase genes (ADH2 and ADH3 on chromosome 4) and one aldehyde dehydrogenase gene (ALDH2 on chromosome 12) exhibit functional polymorphisms. The goal of this study was to determine whether any associations exist between the ADH2, ADH3, and ALDH2 polymorphisms and alcohol dependence in a group of Native Americans. An additional goal was to determine if any associations exist between these polymorphisms and the endophenotype, maximum number of drinks ever consumed in a 24-hour period. METHOD: Mission Indian adults (N=340) were recruited for participation from reservations in southern California. Each participant completed an interview with the Semi-Structured Assessment for the Genetics of Alcoholism. A blood sample was collected from each participant for genotyping at the ALDH2, ADH2, and ADH3 loci. RESULTS: Sixty percent of all participants (72% of men and 53% of women) met lifetime DSM-III-R criteria for alcohol dependence. A significant difference in the ADH2 allele distributions was found between alcohol-dependent and non-alcohol-dependent participants. Those with alcohol dependence were significantly less likely to have the ADH2*3 allele (odds ratio=0.28) and significantly more likely to have the ADH2*1 allele (odds ratio=2.00) than those who were not alcohol dependent. Individuals with ADH2*3 reported a lower number of maximum drinks ever consumed in a 24-hour period, compared to those without this allele. CONCLUSIONS: These results are consistent with genetic linkage studies showing protective associations for alcohol dependence and related behavior on chromosome 4 and suggest that ADH2 polymorphisms may account for these findings. These results also highlight the utility of evaluating protective factors in populations with high rates of alcohol dependence.