Survival of transplanted human corneal stem cells. Case report

PURPOSE: To report a case history involving long-term survival of transplanted human corneal stem cells. METHODS: A male patient with severe bilateral chemical burns received six corneal transplants, all of which failed. He subsequently received combined corneal transplants and stem cell transplants, which have remained clear for 3 and 4 years respectively. One of the donors was female. We studied the gender of the epithelial cells of the cheek of the patient and of the two grafts using fixation and fluorescent in situ hybridization (FISH) analyses. RESULTS: In the graft from the female donor, 30% of the epithelial cells were of female origin. All the epithelial cells from the cheek and the other graft were of male origin. CONCLUSION: This case demonstrates that transplanted human corneal stem cells can survive and replicate in the long-term (3 years) without systemic immunosuppression. The case also indicates that a minority (30%) of healthy transplanted epithelial cells is enough to present a clear graft with a clinically healthy ocular surface.     

Applicability of a modified Edman procedure for measurement of protein adducts: mechanisms of formation and degradation of phenylthiohydantoins

Adducts to N-terminal valine residues in hemoglobin (Hb) are used for monitoring in vivo doses of electrophiles and are quantitated by means of a modified Edman procedure, the "N-alkyl Edman procedure". In the reaction with pentafluorophenyl isothiocyanate, N-alkylated valines cyclize and detach from the protein as pentafluorophenylthiohydantoins (PFPTHs) much more efficiently than do unsubstituted N-terminal valine residues. The mechanisms of this reaction, and of possible degradation reactions, have been studied with model compounds using phenyl- and pentafluorophenyl isothiocyanate. The rapid cyclization to N-alkylvaline-PTHs occurs as a consequence of the influence of substituents on ring formation. This facilitated cyclization favors a direct attack by the thiocarbamoyl nitrogen atom on valine-C-1, and is also observed to occur slowly at unsubstituted N-terminal valines. Such cyclization is favored in protic solvents. Under alkaline conditions and in the presence of air, hydrolytic and oxidative processes give rise to degradation products. The PTH derivatives of N-alkylvaline are less apt to undergo such reactions than are the corresponding derivatives of unsubstituted valine. We conclude that the presence of an N-substituent exerts a greater influence on the cyclization process than the structure of the amino acid or of the Edman reagent. For adducts of different structures, the method has broad applicability, for which the limits, however, are not yet explored. The knowledge from the studies is valid not only for the N-alkyl Edman procedure, but also, to some extent, for the classical Edman degradation reaction. The oxidative side reaction gave rise to the invention of a novel synthesis route for insertion of nucleophiles at carbon-5 in thiohydantoins. The present investigation provides a basis for the N-alkyl Edman procedure, facilitating new toxicological applications.     

Evidence for phosphate adducts in DNA from mice treated with 4-(N-Methyl-N-nitrosamino)-1-(3-pyridyl)-1-butanone (NNK)

The weakly alkylating capacity of phosphotriesters (PTE) has been used for the determination of adducts to phosphate groups in DNA by specific transfer to the strongly nucleophilic compound cob(I)alamin [Cbl(I)]. When enzymatically degraded liver DNA from mice treated with 1-(N-methyl-N-nitrosamino)-4-(3-[3H]pyridyl)-4-oxobutane ([3H]NNK) was added to Cbl(I), a 4-(3-[3H]pyridyl)-4-hydroxy-1-butyl-cobalamin ([3H]PHB-Cbl) complex was formed and determined by HPLC and liquid scintillation counting. The PHB-Cbl formed was compared with a synthetic standard verified by LC/MS and 1H NMR and corresponds to phosphate adducts formed from the pyridyloxobutylating species from NNK and from the pyridylhydroxybutylating species from NNAL, NNK being to a large extent converted to NNAL in vivo. It was concluded that about 22% of the total level of pyridyl (oxo or hydroxy) butyl adducts to DNA was bound to phosphate groups.     

Detection and characterization of a novel functional polymorphism in the GSTT1 gene

A novel functional polymorphism in the GSTT1 gene associated with the non-conjugator phenotype has been identified. Sequencing of GSTT1 cDNA revealed a single nucleotide substitution, 310A>C, that altered the amino acid residue 104 from threonine to proline (T104P). Modelling studies of GSTT1 have suggested that residue 104 is located in the middle of alpha-helix 4. Introduction of an alpha-helix-disrupting proline most likely distorts the conformation of the protein. Individuals that lacked GSTT1 activity and carried the variant allele, tentatively denoted GSTT1*B, had no detectable GSTT1 immunoreactive protein. An allele-specific polymerase chain reaction method was developed to determine the frequency of the GSTT1*B allele. In 497 ethnic Swedes, the frequency of the active GSTT1*A allele was 0.65 [95% confidence interval (CI) 0.62-0.68] whereas the frequencies of the non-functional alleles GSTT1*O and the novel GSTT1*B allele were 0.34 (CI 0.31-0.37) and 0.01 (CI 0.01-0.02), respectively. In 100 Swedish Saamis, the GSTT1*B allele appeared to be slightly more common with a frequency of 0.03 (CI 0.01-0.07). The GSTT1 enzyme activity was measured in erythrocytes using methyl chloride as substrate. Individuals with the GSTT1*A/*A genotype had a two-fold higher GSTT1 activity compared to individuals with the GSTT1*A/*B genotype and subjects with the GSTT1*O/*B genotype totally lacked GSTT1 activity, indicating a strict gene-dose effect. By combining the analyses for the novel single nucleotide polymorphism with analyses for the deletion polymorphism, the accuracy in predicting all three GSTT1 conjugator phenotypes was improved from 96% to 99%.     

Increasing fecal butyrate in ulcerative colitis patients by diet: controlled pilot study

Topical butyrate has been shown to be effective in the treatment of ulcerative colitis (UC). Butyrate is derived from colonic fermentation of dietary fiber, and our aim was to study whether UC patients could safely increase the fecal butyrate level by dietary means. We enrolled 22 patients with quiescent UC (mean age, 44 years; 45% women; median time from last relapse, 1 year) in a controlled pilot trial lasting 3 months. The patients were instructed to add 60 g oat bran (corresponding to 20 g dietary fiber) to the daily diet, mainly as bread slices. Fecal short-chain fatty acids (SCFAs) including butyrate, disease activity, and gastrointestinal symptoms were recorded every 4 weeks. During the oat bran intervention the fecal butyrate concentration increased by 36% at 4 weeks (from 11 +/- 2 (mean +/- SEM) to 15 +/- 2 micromol/g feces) (p < 0.01). The mean butyrate concentration over the entire test period was 14 +/- 1 micromol/g feces (p < 0.05). Remaining fecal SCFA levels were unchanged. No patient showed signs of colitis relapse. Unlike controls, the patients showed no increase in gastrointestinal complaints during the trial. Yet patients reporting abdominal pain and reflux complaints at entry showed significant improvement at 12 weeks that returned to baseline 3 months later. This pilot study shows that patients with quiescent UC can safely take a diet rich in oat bran specifically to increase the fecal butyrate level. This may have clinical implications and warrants studies of the long-term benefits of using oat bran in the maintenance therapy in UC.     

Knowledge of osteoporosis in a Swedish municipality--a prospective study

BACKGROUND: As a part of the Vadstena Osteoporosis Prevention Project, the knowledge of osteoporosis was examined before the intervention program started, after 5 and 10 years. METHODS: At baseline (in 1989) 15% of the population in two Swedish municipalities was randomly invited to the study. The participants in the study group were invited for examination by forearm bone densitometry and a questionnaire concerning lifestyle and risk factors for osteoporosis and also knowledge of osteoporosis, while the subjects in the control group were examined only by questionnaire. Follow-ups were made in 1994 and in 1999. Meanwhile education about osteoporosis was given to the study group, to the public, and to various professionals in the study community. RESULTS: There was a difference in the level of knowledge between the groups prior to the intervention. The rate of increment did not differ significantly between the groups for the study period. Previous participants had 0.58 higher score than new participants in the study group in 1994 (P = 0.031) and 0.76 higher score in 1999 (P < 0.001) regarding the total number of correct answers. The women in the study group had 0.63 higher score than the men in 1994 (P = 0.016) and 1.03 higher score in 1999 (P < 0.001) regarding the total number of correct answers. CONCLUSION: There was no significant effect of a general intervention program concerning the knowledge of osteoporosis in participants in the intervention area compared to the control area.     

Activity level and balance in subjects with mild Alzheimer's disease

To clarify if Alzheimer's disease has an impact on activity level and postural control, we examined 17 elderly diagnosed with mild Alzheimer's disease (MMSE scores 21-29) and 18 age- and gender-matched healthy controls (MMSE scores 27-30) using the Frenchay Activities Index, Bergs Balance Scale, Timed Up & Go and Walking in a Figure of Eight. Mild AD subjects were less active and had lower scores on Bergs Balance Scale, performed Timed Up & Go in longer time and took more steps outside the Figure of Eight, as compared to healthy elderly. This study shows that motor performance is affected already at mild stages of Alzheimer's disease and also that functional performance other than gait may also be impaired.     

Exposure and nasal inflammation in workers heating polyurethane

OBJECTIVE: To test the hypothesis that exposure to thermal-degradation products of polyurethane (PUR), particularly isocyanates, induced nasal inflammation. METHODS: Thirty-eight workers -14 with a history of work-related nasal symptoms (WRS/Nose), and 15 referents without such history - exposed to sprayed and heated PUR glue, were studied with regard to biomarkers of isocyanate exposure [4,4'-diphenylmethane diisocyanate (4,4'-methylenediphenyl diisocyanate; MDI) and 2,4- and 2,6-toluene diisocyanate (TDI), determined as 4,4'-diphenylmethane (U-MDX) and 2,4- and 2,6-toluene (U-2,4 and U-2,6-TDX) diamine in hydrolysed urine and nasal lavage fluid (NAL)], inflammation [albumin; eosinophilic cationic protein (ECP); myeloperoxidase (MPO) and cells in NAL], serum IgG specific for MDI (S-IgG-MDI) and TDI (S-IgG-TDI), and nose symptoms. Nine unexposed office workers were also examined. RESULTS: The exposure to sprayed and heated PUR glue, especially when heated by gun, was associated with the presence of biomarkers of isocyanate exposure in urine; after work the levels [median (range)] in all workers were: U-MDX 0.32 (相似文献