Apoptosis and P53 protein expression after rat spinal cord injury

objective: To determine the DNA damage, neuronal and glial cells apoptosis and expression of P53 protein in the rat spinal cord after compression injury. Methods: Terminal deoxynucleotidyl transferase . mediated dUPT nick end labeling (TUNEL), DNA gel electrophoresis and immunohistochemistry techniques were used to detect DNA fragmentation in the injured rat spinal cord. Results: The apoptosic cells and P53 protein presented at 4 h after spinal cord injury with a maximum presence at 24 h in the injuried T8.9 DNA fragmentation presented typical ladder pattern on agarose gel at 24 h. Conclusion: There are lots of neuronal and glial cells apoptosis with DNA damage after SCl. The P53 protein may play an important role in induc tion of neuronal and glial cells to apoptosis.

Single nucleotide polymorphisms of deoxyribonuclease Ⅰand their expre ssion in Chinese systemic lupus erythematosus patients

Background Previous studies have suggested that interrupted clearance of nuclear DNA-protein complexes after cell death might initiate and propagate systemic lupus erythematosus (SLE). Deoxyribonuclease Ⅰ (DNaseⅠ) may be responsible for the removal of DNA from nuclear antigens at sites of high cell turnover, thus preventing the onset of SLE. The purpose of this study was to genotype the single nucleotide polymorphisms (SNPs) of DNase1 and characterize its gene expression and alternatively spliced transcripts in Chinese patients with SLE in order to unde rstand the pathogenic role of DNase1 in human SLE.Methods Four SNPs located at the 3’ end of the DNase1 gene, as listed on the SNP website, were selected for analysis. Those SNPs with relatively high heterozygosity were chosen for genotyping in 312 Chinese SLE families using the Taqman minor groove binder (MGB) allelic discrimination method. Haplotypes were constructed and linkage disequilibrium tests were performed using GeneHunter. DNase1 mRNA expression was detected using real-time polymerase chain reaction (PCR), and alternatively spliced transcripts were isolated using capillary electrophoresis. Any effects the specific SNP haplotypes had on DNase1 gene expression and the alternatively spliced transcripts were also assessed.Results rs179982 and rs1053874 had high heterozygosity, about 0.5 in this Chinese cohort, while rs1059857 was also found to be heterozygous. Analysis of the haplotype combining rs179982-rs1030874 (C-G) and rs179982-rs1030874-rs1059857 (C-G-G) revealed a skewed transmission in favor of affected offspring. DNase1 gene expression was higher in SLE patients than in normal controls (P<0.001), but this was not related to disease activity or SNP haplotype. Capillary electrophoresis revealed that the pattern of alternatively spliced transcripts in patients differed from that of normal controls. Furthermore, different SNP haplotype combinations generated different transcript patterns in SLE patients. Conclusions The SNP haplotypes are in linkage disequilibrium in Chinese SLE patients and may induce the disease through a modification of DNase1 mRNA splicing rather than at the level of mRNA expression. There is a relatively unique transcript band in SLE patients independent of special haplotype, which suggests that other unknown factors might be involved in adjusting gene expression.

Traumatic injury is associated with reduced deoxyribonuclease activity and dysregulation of the actin scavenging system

Background:Traumatic injury is associated with increased concentrations of cell-free DNA(cfDNA)in the circulation,which contribute to post-injury complications.The endonuclease deoxyribonuclease 1(DNase-1)is responsible for removing 90%of circulating cfDNA.Recently,DNase activity was reported to be significantly reduced following major non-traumatic brain injury(TBI),but the processes responsible were not investigated.Moreover,it is not known how quickly following injury DNase activity is reduced and whether this also occurs after TBI.Methods:At 3 post-injury time points(≤1,4–12 and 48–72 hours),blood samples were obtained from 155 adult trauma patients that had sustained an isolated TBI(n=21),TBI with accompanying extracranial injury(TBI^(+))(n=53)or an extracranial injury only(ECI)(n=81).In addition to measuring cfDNA levels and the activity and expression of DNase,circulating concentrations of monomeric globular action(G-actin),an inhibitor of DNase-1,and the actin scavenging proteins gelsolin(GSN)and vitamin D binding protein(VDBP)were determined and values compared to a cohort of healthy controls.Results:Significantly elevated concentrations of plasma cfDNA were seen in TBI,TBI^(+)and ECI patients at all study time points when compared to healthy controls.cfDNA levels were significantly higher at≤1 hour post-injury in ECI patients who subsequently developed multiple organ dysfunction syndrome when compared to those who did not.Plasma DNase-1 protein was significantly elevated in all patient groups at all sampling time points.In contrast,DNase enzyme activity was significantly reduced,with this impaired function evident in TBI^(+)patients within minutes of injury.Circulating concentrations of G-actin were elevated in all patient cohorts in the immediate aftermath of injury and this was accompanied by a significant reduction in the levels of GSN and VDBP.Conclusions:The post-traumatic increase in circulating cfDNA that occurs following extracranial trauma and TBI is accompanied by reduced DNase activity.We propose that,secondary to reduced GSN and VDBP levels,elevated circulating concentrations of G-actin underlie the post-injury reduction in DNase activity.Reducing circulating cfDNA levels via therapeutic restoration of DNase-1 activity may improve clinical outcomes post-injury.