Reduction of scar formation in full-thickness wounds with topical celecoxib treatment

Adult wound repair occurs with an initial inflammatory response, reepithelialization, and the formation of a permanent scar. Although the inflammatory phase is often considered a necessity for successful adult wound healing, fetal healing studies have shown the ability to regenerate skin and to heal wounds in a scarless manner in the absence of inflammation. The cyclooxygenase-2 (COX-2) enzyme, a known mediator of inflammation, has been shown to contribute to a variety of inflammatory conditions and to the development of cancer in many organs. To examine the role of COX-2 in the wound healing process, incisional wounds were treated topically with the anti-inflammatory COX-2 inhibitor celecoxib. Acutely, celecoxib inhibited several parameters of inflammation in the wound site. This decrease in the early inflammatory phase of wound healing had a significant effect on later events in the wound healing process, namely a reduction in scar tissue formation, without disrupting reepithelialization or decreasing tensile strength. Our data suggest that in the absence of infection, adult wound healing is able to commence with decreased inflammation and that anti-inflammatory drugs may be used to improve the outcome of the repair process in the skin by limiting scar formation.     

Categorization and deductive reasoning: predictors of instrumental activities of daily living performance in adults with brain injury.

OBJECTIVE: The purpose of this study was to examine the predictive relationship of categorization and deductive reasoning abilities to performance of instrumental activities of daily living (IADL) among adults with brain injury. METHOD: A correlational research design was used to examine the performance of 19 patients from a neurosurgery acute care unit on Toglia's Category Assessment (TCA), the Deductive Reasoning test (DR), and the Observed Tasks of Daily Living-Revised. RESULTS: Multiple regression analysis indicated that categorization and deductive reasoning accounted for a significant proportion of the variance in IADL performance scores, with deductive reasoning making the greater contribution. Demographic characteristics such as age and education reduced the power of the previous variables, especially of the categorization. CONCLUSION: The results of this study indicate that assessments of categorization and deductive reasoning abilities of persons with brain injury may be good predictors of IADL functional performance. Results support the use of the TCA and DR as quick measures of a person's cognitive and functional abilities. Furthermore, this study provides preliminary results to support that categorization and deductive reasoning abilities are fundamental for the performance of IADL skills.     

Analysis of gene expression in MOG-induced experimental autoimmune encephalomyelitis after treatment with a novel brain-penetrating antioxidant

Accumulating data from experimental studies indicate that oxidative stress has a major role in the pathogenesis of multiple sclerosis (MS). It has been suggested that local production of reactive oxygen species, probably by macrophages, mediates axonal damage in both MS patients and the mouse model experimental autoimmune encephalomyelitis (EAE). We have shown previously that our novel brain-penetrating antioxidant, N-acetylcysteine amide (AD4), reduces the clinical and pathological symptoms, including inflammation and axonal damage in myelin oligodendrocyte glycoprotein (MOG)-induced chronic EAE in mice. The aim of this study was to examine the molecular mechanism by which AD4 exerts protection in MOG-induced EAE mice. Therefore, we analyzed gene-expression profile in the spinal cords of MOG-induced chronic EAE mice and compared them with MOG-induced mice treated with AD4, using a cDNA microarray. We found that MOG treatment up-regulated genes encoding growth factors, cytokines, death receptors, proteases, and myelin structure proteins, whereas MOG- and AD4-treated mice demonstrated gene expression profiles similar to that seen in na?ve healthy mice. In conclusion, our study shows that chronic AD4 administration suppresses the induction of various pathological pathways that play a role in EAE and probably in MS.     

Effects of MK-801 and Phenytoin on Flurothyl-Induced Seizures During Development

Summary We determined the effects of the N-methyl-Daspartate (NMDA) receptor blocker MK-801 (0.05, 0.1, and 0.5 mg/kg intraperitoneally, i.p.) and phenytoin (PHT, 5, 10, and 20 mg/kg i.p.) on flurothyl-induced clonic and tonic-clonic seizures in 9-, 1 5, 30-, and 60-day-old male rats. Both agents had seizure-, age-, and dose-specific effects. The highest dose of MK-801 was anticonvulsant against clonic flurothyl-induced seizures only in 9- and 60-day-old rats, but suppressed tonic-clonic seizures in all ages. The lowest dose of MK-801 (0.05 mg/kg) produced significant anticonvulsant effects only in 15 day old rats. PHT did not have any effect on clonic seizures throughout development. Both doses of PHT (10 and 20 mg/kg) were anticonvulsant against tonic-clonic seizures in adult rats but not in any other age group. The results indicate that NMDA receptors play an important role in tonic-clonic flurothyl-induced seizures throughout development (especially in 15-day-old rats) and that the anticonvulsant effects of PHT may vary at different stages of brain development.     

Cellular and molecular characteristics of an immortalized ataxia-telangiectasia (group AB) cell line

Ataxia-telangiectasia (A-T) is a multisystem hereditary disease featuring neurodegeneration, immunodeficiency, extreme cancer proneness, chromosomal instability, and radiosensitivity. A-T is found in many ethnic groups, and is genetically heterogeneous: four complementation groups have been identified in A-T so far. Attempts to isolate the A-T gene are based in part on gene transfer experiments, using permanent A-T fibroblast lines, obtained by transformation with SV40. "Immortalization" of A-T primary diploid fibroblasts using SV40 is difficult, possibly because of the chromosomal instability of these cells. The number of currently available permanent A-T fibroblast lines is small, and not all of them have been assigned to specific complementation groups. Using the assay of X-ray induced inhibition of DNA synthesis, we have assigned the A-T strain AT22IJE to complementation group AB. Origin-defective SV40 was used to transfect these cells, and one transformant (AT22IJE-T), which survived crisis, was found to have the typical characteristics of permanent cell lines obtained in this way. "In-gel renaturation" analysis did not show any DNA amplification of high degree in AT22IJE-T. Cytogenetic analysis showed considerable chromosomal instability in the new cell line, and medium conditioned by these cells contained the clastogenic activity which is characteristic of the parental strain as well. Other parameters of the "cellular A-T phenotype" have also been retained in the immortalized cells: hypersensitivity to the lethal effects of X-rays and neocarzinostatin, as well as "radioresistant" DNA synthesis. However, the sensitivity of AT22IJE-T to both DNA-damaging agents is less pronounced than that of the parental cells. The capacity of the cells for uptake of foreign DNA was tested by introducing into them the plasmid pRSVneo, using three different transfection methods. Satisfactory frequency of G418-resistant transfectants (0.66%) was achieved using a protocol recently published by Chen and Okayama (Mol. Cell Biol., 7: 2745-2752, 1987), which was found to be superior to the traditional calcium phosphate transfection method and to the polybrene-based method.     

Polyl-histidine

Poly-l-histidine (PHSTD) of molecular weight 26,000 induced the generation of large amounts of Superoxide (O ₂ ^– ) and hydrogen peroxide (H₂O₂) in human neutrophils (PMNs). Despite its low solubility at neutral pH, PHSTD was bound very rapidly to the PMN surfaces. Maximal generation of O ₂ ^– took place with 4–5 ×10^–6 M of PHSTD, starting after a lag of about 25 sec and proceeding for 15–17 min at a rate of 150 nmol/10⁷ PMNs/min, suggesting that this polycation is one of the most potent stimulators of O ₂ ^– generation known, PHSTD was found to be non-toxic for PMNs even at millimolar concentrations. Generation of O ₂ ^– by PHSTD depended on extracellular calcium; it was inhibited by calcium channel blockers and by trifluoperazine, and it triggered a sharp rise in intracellular calcium as determined by the Quin 2 fluorescence technique. The generation of both O ₂ ^– and H₂O₂ by PHSTD was partially inhibited by cytochalasin B or (CYB, CYE). On the other hand, CYB markedly enhanced the generation of both O ₂ ^– and H₂O₂ following stimulation of PMNs either by PHSTD, polyarginine, histone, or by antibody-opsonized group A streptococci. Electron microscopic analysis and NBT reduction tests revealed that both PHSTD and PHSTD-opsonized streptococci were avidly phagocytosed by PMNs. Since CYB totally inhibited internalization of both PHSTD and the PHSTD-opsonized streptococci, it was suggested that these agents stimulated oxygen radical generation mainly on the leukocyte surfaces. Complexes (CX) formed between PHSTD and polyanethole sulfonate (a strong polyanion) or between histone and the polyanion mimicked immune CX in their ability to trigger the generation of large amounts of O ₂ ^– which were inhibited by CYB. Generation of O ₂ ^– and chemiluminescence either by PHSTD or by PHSTD-opsonized streptococci were markedly inhibited by poly-l-glutamate, suggesting that PHSTD acted as a cationic agent which interacted via electrostatic forces with some negatively charged sites in the leukocyte membrane. Generation of H₂O₂ by PHSTD was also markedly inhibited by deoxyglucose, KCN, DASA, as well as by the lipoxygenase inhibitors nordihydroguaiaretic acid, phenidone, and propylgallate. On the other hand, cyclooxygenase inhibitors such as aspirin, indomethacin, and piroxicam were inactive, suggesting that arachidonic acid metabolism via lipoxygenase pathway might have been involved in the activation by PHSTD of the NADPH oxidase in PMNs. PHSTD may mimic the effects of antibodies both as an opsonin and as a potent stimulator of the respiratory burst in PMNs and may thus serve as a model for further study of leukocyte-bacteria interactions in infectious and inflammatory sites and of the pathogenicity of immune complexes.Supported by a research grant from Dr. S. M. Robbins of Cleveland, Ohio.     

PCR detection of amplified 132 bp repeats in Marek's disease virus type 1 (MDV-1) DNA can serve as an indicator for critical genomic rearrangement leading to the attenuation of virus virulence

A radioactive PCR test was developed that amplified the very virulent Marek's disease virus-1 (vvMDV-1) DNA sequence containing the 132 bp repeats. In apathogenic MDV-1 (CVI 988, Rispens), amplified DNA bands containing multiple copies of 132 bp repeats were identified. In the present study this PCR technique was used to monitor the passage level of vvMDV-1 in chicken embryo fibroblasts (CEF) in which the number of tandem 132 bp repeats was increased. It was found that at passage level 32 of vvMDV-1-B isolate, the 132 bp tandem repeat was already markedly amplified and its pattern resembled that of the MDV-1 (CVI 988, Rispens) vaccine virus DNA. In the vvMDV-1Z strain, amplification of the 132 bp repeat was not detectable at a similar passage level. The PCR test demonstrated that the apathogenic MDV-1 Md11/75c virus developed by extensive in vitro passaging has amplified 132 bp DNA repeats similar to those of the commercial vaccine virus (CVI 988, Rispense). It was also found that the pattern of viral RNA from infected cells detectable by Northern blot hybridization was markedly changed from a 2.4 kb RNA species in cells infected with vvMDV-1 viruses, to four RNA species (ranging from 2.2 to 4.4 kb) in cells infected with passage 32 of MDV-1-B strain, to a very large number of undefined RNA species synthesized in cells infected with attenuated MDV-1 viruses (CVI 988, Rispens and Md 11/75c).     

Identification and characterization of linear B-cell epitopes of beta-globulin, a major allergen of sesame seeds

BACKGROUND: The increased consumption of foods containing sesame seeds is paralleled by an increase in reported sesame-induced allergic reactions. OBJECTIVE: This study aimed at identifying and characterizing the linear B-cell epitopes of the 14-kd beta-globulin, the major allergen of sesame seed. METHODS: A peptide containing 71 amino acids (peptide B) was previously identified by us as the IgE-binding region on beta-globulin. To determine the amino acid sequence of the IgE-binding sites on peptide B, we synthesized overlapping peptides 20 and 10 amino acid residues long that span the entire length of peptide B, which were offset from each other by 10 and 2 amino acid residues, respectively. Sera from 20 subjects given diagnoses of allergy to sesame beta-globulin served to identify the epitopes by using the dot-blot test. RESULTS: At least 9 different IgE-recognition sites were identified on peptide B. Three of them, numbers 2, 3, and 13 (corresponding to amino acids 46-55, 48-57, and 76-86, respectively, in the beta-globulin sequence), appeared to be immunodominant IgE-binding epitopes. Also, these peptides were best recognized in terms of intensity of response. There was no obvious sequence motif shared by the 9 different IgE-binding epitopes of beta-globulin. However, approximately 60% of the amino acids represented in the epitopes are hydrophobic residues. CONCLUSION: Identification of the IgE-binding epitopes might provide a better understanding of the functional role the allergens play in the disease and might have implications for immunodiagnosis and probably immunotherapy.     

A novel point mutation (I137T) in the conserved 5-phosphoribosyl-1-pyrophosphate binding motif of hypoxanthine-guanine phosphoribosyltransferase (HPRTJerusalem) in a variant of Lesch-Nyhan syndrome

We identified a novel point mutation (I137T) in the hypoxanthine-guanine phosphoribosyltransferase (HPRT; EC 2.4.2.8) encoding gene, in a patient with partial deficiency of the enzyme (variant of Lesch-Nyhan syndrome). The mutation, ATT to ACT, resulting in substitution of isoleucine to threonine, occurred at codon 137 (exon 6), which is within the region encoding the binding site for 5-phosphoribosyl-1-pyrophosphate (PRPP). We suggest the mechanism by which the mutation-induced structural alteration of HPRT reduced the affinity of the enzyme for PRPP.