[Screening of pathogenic molecular markers of Staphylococcus aureus in children based on whole genome sequencing technology]. / åŸºäºŽå ¨åŸºå› ç»„æµ‹åºæŠ€æœ¯ç­›é€‰å„¿ç«¥é‡‘é»„è‰²è‘¡è„çƒèŒçš„è‡´ç— ç›¸å ³åˆ†å­æ ‡å¿—ç‰©.

OBJECTIVES: To explore the molecular characteristics of Staphylococcus aureus (S. aureus) in children, and to compare the molecular characteristics of different types of strains (infection and colonization strains) so as to reveal pathogenic molecular markers of S. aureus. METHODS: A cross-sectional study design was used to conduct nasopharyngeal swab sampling from healthy children in the community and clinical samples from infected children in the hospital. Whole genome sequencing was used to detect antibiotic resistance genes and virulence genes. A random forest method to used to screen pathogenic markers. RESULTS: A total of 512 S. aureus strains were detected, including 272 infection strains and 240 colonization strains. For virulence genes, the carrying rates of enterotoxin genes (seb and sep), extracellular enzyme coding genes (splA, splB, splE and edinC), leukocytotoxin genes (lukD, lukE, lukF-PV and lukS-PV) and epidermal exfoliating genes (eta and etb) in infection strains were higher than those in colonization strains. But the carrying rates of enterotoxin genes (sec, sec3, seg, seh, sei, sel, sem, sen, seo and seu) were lower in infection strains than in colonization strains (P<0.05). For antibiotic resistance genes, the carrying rates of lnuA, lnuG, aadD, tetK and dfrG were significantly higher in infection strains than in colonization strains (P<0.05). The accuracy of cross-validation of the random forest model for screening pathogenic markers of S. aureus before and after screening was 69% and 68%, respectively, and the area under the curve was 0.75 and 0.70, respectively. The random forest model finally screened out 16 pathogenic markers (sem, etb, splE, sep, ser, mecA, lnuA, sea, blaZ, cat(pC233), blaTEm-1A, aph(3')-III, ermB, ermA, ant(9)-Ia and ant(6)-Ia). The top five variables in the variable importance ranking were sem (OR=0.40), etb (OR=3.95), splE (OR=1.68), sep (OR=3.97), and ser (OR=1.68). CONCLUSIONS: The random forest model can screen out pathogenic markers of S. aureus and exhibits a superior predictive performance, providing genetic evidence for tracing highly pathogenic S. aureus and conducting precise targeted interventions.

[Dose-response relationship between age and Streptococcus pneumoniae vaccination coverage in kindergarten children]. / 幼儿园儿童年龄与肺炎链球菌疫苗覆盖率间的剂量-ååº”å ³ç³».

OBJECTIVES: To investigate the potential relationship between age and Streptococcus pneumoniae vaccination coverage in kindergarten children, and to provide a basis for guiding vaccination and developing new protein vaccines. METHODS: The stratified cluster random sampling method was used to select 1 830 healthy children from six kindergartens in Shunde District, Foshan City, China, and nasopharyngeal swabs were collected for the isolation and identification of Streptococcus pneumoniae. The logistic regression model based on restricted cubic spline was used to analyze the dose-response relationship between age and Streptococcus pneumoniae vaccination coverage. RESULTS: The rate of nasal Streptococcus pneumoniae carriage was 22.46% (411/1 830) among the kindergarten children, with the predominant serotypes of 6B, 19F, 15A, 23A, 34, and 23F. The coverage rates of 10-valent pneumococcal conjugate vaccine (PCV10) and 13-valent pneumococcal conjugate vaccine (PCV13) were 53.0% and 57.9%, respectively, and there was a significant non-linear dose-response relationship between age and the coverage rates of PCV10 and PCV13 (P<0.05), with a higher coverage rate of PCV10 (88.0%) and PCV13 (91.1%) in the children aged 2 years. There was a significant non-linear dose-response relationship between age and the coverage rates of pilus islet 1 (PI-1) and pilus islet 2 (PI-2) (P<0.05), with a lower vaccination coverage rate for PI-1 (37.7%) and PI-2 (16.1%). The coverage rates of PI-1 (13.0%-58.5%) and PI-2 (6.0%-29.4%) were lower in all age groups. The virulence genes lytA (99.5%) and ply (99.0%) associated with candidate protein vaccines showed higher vaccination coverage rates. CONCLUSIONS: There is a significant non-linear dose-response relationship between the age of kindergarten children and the coverage rates of PCV10 and PCV13 serotypes, and kindergarten children aged 2 years have a relatively high coverage rate of PCV. The high prevalence of the virulence genes lytA and ply shows that they are expected to become candidate virulence factors for the development of a new generation of recombinant protein vaccines.

Co-carriage of Streptococcus pneumoniae and Moraxella catarrhalis among preschool children and its influencing factors. / å­¦é¾„å‰å„¿ç«¥è‚ºç‚Žé“¾çƒèŒå’Œå¡ä»–èŽ«æ‹‰èŒæºå¸¦çŠ¶æ€åŠå½±å“å› ç´ åˆ†æž.

OBJECTIVES: To investigate the carriage status of Streptococcus pneumoniae (S.pneumoniae) and Moraxella catarrhalis (M. catarrhalis) in preschool children and the influencing factors for the carriage status. METHODS: The stratified cluster sampling method was used to select 2 031 healthy children from seven kindergartens in Shunde District of Foshan in Guangdong, China. Nasal swabs were collected from all children for the isolation and identification of S. pneumoniae and M. catarrhalis. The carriage status of S. pneumoniae and M. catarrhalis was analyzed in terms of its association with demographic features and hospital- and community-related factors. RESULTS: The carriage rates of S. pneumoniae and M. catarrhalis were 21.81% and 52.44%, respectively among the children. The co-carriage rate of S. pneumoniae and M. catarrhalis was 14.87%. The correspondence analysis showed that the factors such as lower grade, non-local registered residence, living in rural areas, small living area, history of respiratory tract infection but no history of antibiotic use, allergic skin diseases, and no hospital-related exposure history were significantly associated with the co-carriage of S. pneumoniae and M. catarrhalis among the children (P<0.05). CONCLUSIONS: Co-carriage of S. pneumoniae and M. catarrhalis can be observed in preschool children. Young age, poor living environment, a history of respiratory tract infection but no history of antibiotic use, allergic skin diseases, and no hospital-related exposure history are important risk factors for the co-carriage of S. pneumoniae and M. catarrhalis in preschool children.

Effects of norepinephrine on microglial neuroinflammation and neuropathic pain.

Norepinephrine (NE) is an important neurotransmitter in the central nervous system. NE is released from locus coeruleus neurons and is involved in a variety of physiological and pathological processes. Neuroinflammation is a common manifestation of many kinds of neurological diseases. The activation of microglia directly affects the status of neuroinflammation. Several kinds of adrenergic receptors, which anchor on microglia and can be regulated by NE, affect the activation of microglia and neuroinflammation. NE influences chronic pain, anxiety, and depression by regulating the activation of microglia.

[Concentrations and Distribution of Metals in the Core Sediments from Estuary and City Section of Liaohe River].

The particle size, total organic carbon (TOC), total nitrogen (TN), C/N ratio and metal concentrations as well as activities of 210Pb were determined in Liaohe River estuary area (LN-2) and Shenyang area (LN-5), and the organic matter resources were discussed in two core sediments. Also the index of geoaccumulation (Igeo) and enrichment factors (EFs) methods were applied to evaluate the state of heavy metal contamination in the studied sties. The study showed that both sediment cores LN-2 and LN-5 were dominated by silts, and the vertical variations of grain-size composition and organic matter were well distributed in LN- 2 while fluctuated in LN-5. According to the organic matter source analysis through C/N ratio, C/N ratio varied in the scale of 5. 24-7.93 in LN-2 which was dominated by river source, and 9.94-14.21 in LN-5 which was dominated by terrestrial input. Al, Ca, Fe, Mn, Ni, Cu, Zn, Cd, Pb and Cr in two sediment cores had different vertical changing rules, Ni and Zn in LN-2 as well as Pb and Zn in LN-5 were affected by both natural and human factors, other elements had similar distributions to those of organic matters, which showed that these elements were mainly affected by the natural activities. Based on Igeo and EFs, both sediment cores were more severely polluted with Ni, Zn and Pb than other metals. The effects of human activities on the environment were also discussed in this study, combined with the economical development of Liaoning Province and the studied sites in the past 20 years.

[Relationship between genotypes and clinical phenotypes in patients from 7 hemophilia A families].

OBJECTIVE: To study the mutation types of factor VIII (FVIII) gene in patients from 7 hemophilia A (HA) families and the relationship between FVIII gene mutations and clinical phenotypes. METHODS: A total of 8 patients from 7 HA families were recruited. The activated partial thromboplastin time (APTT) and factor VIII coagulant activity (VIII:C) in these patients were measured. Polymerase chain reaction (PCR) was performed to analyze FVIII gene intron 1 and 22 inversions. For patients without the FVIII intron inversions, direct sequencing was performed to determine their mutation types and other related members of their families were also tested by PCR and sequencing to analyze the corresponding mutation sites. RESULTS: The ranges of APTT and VIII:C of the 8 patients were 91.6-131 seconds and 0.8%-2%, respectively. FVIII gene intron 22 inversion was not detected, while intron 1 inversion was detected in one patient. There were 5 types of mutations in FVIII gene detected in the remaining 7 patients, including 6 patients with mutations in exon 14 and 1 patient with mutation in exon 23; all of the 5 types of mutations were single nucleotide mutations. Among the detected mutations in FVIII gene, p.His1202LeufsX16 (c.3666delA) detected in one patient was found to be a previously unreported mutation in FVIII gene. CONCLUSIONS: FVIII gene exon 14 is a hot-spot mutation region and p.His1202LeufsX16 is found to be a novel mutation in FVIII gene.

Development of aquatic life criteria for triclosan and comparison of the sensitivity between native and non-native species.

Triclosan (TCS) is an antimicrobial agent which is used as a broad-spectrum bacteriostatic and found in personal care products, and due to this it is widely spread in the aquatic environment. However, there is no paper dealing with the aquatic life criteria of TCS, mainly result from the shortage of toxicity data of different taxonomic levels. In the present study, toxicity data were obtained from 9 acute toxicity tests and 3 chronic toxicity tests using 9 Chinese native aquatic species from different taxonomic levels, and the aquatic life criteria was derived using 3 methods. Furthermore, differences of species sensitivity distributions (SSD) between native and non-native species were compared. Among the tested species, demersal fish Misgurnus anguillicaudatus was the most sensitive species, and the fishes were more sensitive than the aquatic invertebrates of Annelid and insect, and the insect was the least sensitive species. The comparison showed that there was no significant difference between SSDs constructed from native and non-native taxa. Finally, a criterion maximum concentration of 0.009 mg/L and a criterion continuous concentration of 0.002 mg/L were developed based on different taxa, according to the U.S. Environmental Protection Agency guidelines.

[Drug resistance of methicillin resistant Staphylococcus strains in burn ward and relative analysis].

OBJECTIVE: To study the distribution and drug resistance of methicillin resistant Staphylococcus strains in various specimens of inpatients in burn wards, and to provide reference for clinical treatment. METHODS: Bacteria were isolated from specimens of wound exudate, blood, sputum, and bronchoalveolar lavage fluid etc., which were collected from patients hospitalized in our burn wards from January 2008 to December 2010. The bacteria were routinely cultured and identified. Drug resistance of the Staphylococci to 15 antibiotics commonly used in clinic was identified by K-B disk diffusion method. Data were processed with statistical software WHONET 5.5. The homology of 40 strains of methicillin resistant Staphylococcus aureus (MRSA) was analyzed by pulsed-field gel electrophoresis (PFGE). RESULTS: Altogether 386 strains of Staphylococcus were isolated, including 196 strains of Staphylococcus aureus and 190 strains of coagulase negative Staphylococcus. The mean annual isolation rates of MRSA and methicillin resistant coagulase negative Staphylococcus (MRCoNS) were respectively 73.00% (143/196) and 74.20% (141/190). The resistance rates of MRSA and MRCoNS to ß-lactams drugs, such as penicillin, oxacillin, cefazolin, and cefuroxime were 100.00% in every year. No Staphylococcus strains resistant to vancomycin, teicoplanin, or linezolid were found. Three different PFGE patterns A, B, and C were identified among 40 MRSA strains, including 33 strains of type A (30 strains in sub-type A1 and 3 strains in sub-type A2), 6 strains of type B (respectively 3 strains in sub-types B1 and B2), and 1 strain of type C. CONCLUSIONS: The isolation rates of MRSA and MRCoNS were high in our burn wards from January 2008 to December 2010. All of them showed strong drug resistance property, and they were multidrug resistant. The most prevalent strain was PFGE type A.

[Application of STR genetic marker system in the detection of hemophilia A carriers in Guangxi, China].

OBJECTIVE: To establish a fast and simple genetic diagnosis technique based on a reliable, short tandem repeat (STR) genetic marker system for the detection of hemophilia A carriers in Guangxi, China. METHODS: Fluorescent PCR and capillary electrophoresis were used for allele genotyping at three intragenic/extragenic STR loci (F8Int13, DXS1073, and DXS9901) of FVIII gene in the members of 10 hemophilia A families in Guangxi, so as to evaluate the diagnostic efficiency of the STR genetic marker system for detection of hemophilia A carriers. Then the STR genetic marker system was used to detect hemophilia A carriers among examinees. RESULTS: In the 10 hemophilia A families, 11 confirmed female carriers had the same allele fragment lengths at the three STR loci (F8Int13, DXS1073, and DXS9901) as the probands. Of the 8 females examined, 5 had allele fragments at the three STR loci (F8Int13, DXS1073, and DXS9901) which were identical to those of the probands, and thus they were diagnosed as hemophilia A carriers. CONCLUSIONS: Genetic analysis at the three STR loci (F8Int13, DXS1073, and DXS9901) can be used to detect hemophilia A carriers rapidly and provide reliable basis for prenatal diagnosis of hemophilia A.

[Spatial distribution and risk assessment of polycyclic aromatic hydrocarbons in partial surface sediments of Liaohe River].

Nineteen surface sediment samples were collected from Liaohe River and analysed for polycyclic aromatic hydrocarbons (PAHs) with GC-MS. The concentrations of 15 detected PAH compounds ranged from 123.5 to 21 233.4 ng x g(-1) in dry season with the average concentration was 3208.1 ng x g(-1); and were 37.9 to 9014.0 ng x g(-1) in wet season with the average concentration was 1612.0 ng x g(-1). The isomeric ratio method using characteristic compounds of PAHs for source analysis, and the main source was attributed to fuel combustion. Average commercial use of sediment quality benchmarks (mSQG-Q) on the PAHs risk assessment of Liaohe Basin was applied, fluorene and pyrene showed the potential of low-level ecological effects. In wet season, Fushun segment L3-1 points have a higher ecological risk. In dry season, Shenyang segment L1-1 point, Fushun segment L3-1 point and L3-2 point pose a strong negative effect.

[Molecular analysis of two pedigrees with inherited coagulation factor VII deficiency].

OBJECTIVE: To analyze the gene mutation in two pedigrees of inherited coagulation factor VII (FVII) deficiency, and investigate the relationship between the genotype and phenotype. METHOD: The coagulation function and coagulation factors activity of probands were detected for phenotype diagnosis, all exons and junctions of FVII gene from the family members' genomic DNA were amplified using polymerase chain reaction (PCR), and detected the gene mutation by direct sequencing. Mutations were confirmed by reverse sequencing. RESULT: The prothrombin time (PT) of proband 1 was 265.2 s, FVII:C was 22% and the PT of proband 2 was > 120 s, FVII:C was 1%. Homozygous 17844G→A mutation in No. 8 exon of FVII gene was identified in the proband 1 resulting in Gly343Ser, and heterozygosity for the same mutations were confirmed in his parents and a sister. The proband 2 was compound heterozygous, one mutation was the same as the proband 1 but was a heterozygosity that can also found in his mother and brother; the other heterozygosity mutation was located on No. 8 exon 18055G→A that resulted in Gln413Arg which was inherited from his father. CONCLUSION: No. 8 exon of FVII gene encodes catalytic domain. Mutation found in those domain could change the FVII catalytic domain spatial structure, affected FVII function and stability, and the sufferer of homozygote and compound heterozygous may have clinical bleeding tendency. Almost no clinical findings in simple heterozygotes, however, a few of heterozygotes could have a tendency of bleeding because of genetic polymorphism which would reduce the FVII:C.

Early life seizures cause long-standing impairment of the hippocampal map.

Children with seizures are at risk for long-term cognitive deficits. Similarly, recurrent seizures in developing rats are associated with deficits in spatial learning and memory. However, the pathophysiological bases for these deficits are not known. Hippocampal place cells, cells that are activated selectively when an animal moves through a particular location in space, provides the animal with a spatial map. We hypothesized that seizure-induced impairment in spatial learning is a consequence of the rat's inability to form accurate and stable hippocampal maps. To directly address the cellular concomitants of spatial memory impairment, we recorded the activity of place cells from hippocampal subfield CA1 in freely moving rats subjected to 100 brief flurothyl-induced seizures during the first weeks of life and then tested them in the Morris water maze and radial-arm water maze followed by place cell testing. Compared to controls, rats with recurrent seizures had marked impairment in Morris water maze and radial-arm water maze. In parallel, there were substantial deficits in action potential firing characteristics of place cells with two major defects: i) the coherence, information content, center firing rate, and field size were reduced compared to control cells; and ii) the fields were less stable than those in control place cells. These results show that recurrent seizures during early development are associated with significant impairment in spatial learning and that these deficits are paralleled by deficits in the hippocampal map. This study thus provides a cellular correlate for how recurrent seizures during early development lead to cognitive impairment.

Cognitive dysfunction after experimental febrile seizures.

While the majority of children with febrile seizures have an excellent prognosis, a small percentage are later discovered to have cognitive impairment. Whether the febrile seizures produce the cognitive deficits or the febrile seizures are a marker or the result of underlying brain pathology is not clear from the clinical literature. We evaluated hippocampal and prefrontal cortex function in adult rats with a prior history of experimental febrile seizures as rat pups. All of the rat pups had MRI brain scans following the seizures. Rats subjected to experimental febrile seizures were found to have moderate deficits in working and reference memory and strategy shifting in the Morris water maze test. A possible basis for these hippocampal deficits involved abnormal firing rate and poor stability of hippocampal CA1 place cells, neurons involved in encoding and retrieval of spatial information. Additional derangements of interneuron firing in the CA1 hippocampal circuit suggested a complex network dysfunction in the rats. MRI T2 values in the hippocampus were significantly elevated in 50% of seizure-experiencing rats. Learning and memory functions of these T2-positive rats were significantly worse than those of T2-negative cohorts and of controls. We conclude that cognitive dysfunction involving the hippocampus and prefrontal cortex networks occur following experimental febrile seizures and that the MRI provides a potential biomarker for hippocampal deficits in a model of prolonged human febrile seizures.

Effect of topiramate on cognitive function and single units from hippocampal place cells following status epilepticus.

Topiramate, an antiepileptic drug with multiple mechanisms of action, was assessed as a neuroprotective agent following status epilepticus. We administered topiramate or normal saline chronically beginning 1 hour after cessation of lithium pilocarpine-induced status epilepticus. Control animals not subjected to status epilepticus were also treated with topiramate or normal saline. Following completion of the topiramate treatment, animals were tested in the water maze to assess spatial learning and underwent in vivo single-cell place cell recordings. Spontaneous seizure frequency following status epilepticus in the topiramate-treated rats was similar to that in the rats treated with saline. Following status epilepticus, rats had profound deficits in water maze performance and place cell function. Rats subjected to status epilepticus and treated with topiramate were also severely impaired in the water maze, but performed slightly better than rats treated with saline. Following status epilepticus, topiramate-treated rats did not differ from rats treated with normal saline in the platform switch, a test of prefrontal function. Although place cell firing patterns were similar in both the topiramate- and saline-treated rats, rats treated with topiramate had higher information content scores than rats treated with saline. Topiramate-treated animals had less supragranular sprouting following status epilepticus than nontreated rats. Control animals treated with topiramate did not differ from saline-treated controls on any measures. Taken together, this study shows that topiramate administered following status epilepticus has modest neuroprotective effects.

[Effects of glycine on apoptosis in murine cardiomyocyte suffering from ischemia and hypoxia].

OBJECTIVE: To investigate the effects of glycine on apoptosis in murine cardiomyocyte suffering from ischemia and hypoxia. METHODS: The primary passage of cultured cardiomyocytes from neonatal rats were subjected to ischemia and hypoxia, and the cells were divided into IH (without other treatment), and G (with treatment of 5 mmol/L glycine) groups. Normal murine cardiomyocytes served as control (C group). Cardiomyocytes were cultured for 6 hours in vitro. Apoptosis, mitochondrial membrane potential and its distribution, the condition of mitochondria permeability transition pore (mPTP) were observed with expression of fluorescence intensity. The activity of caspase-3 was observed by Laser Scanning staining. RESULTS: (1) Apoptosis: the fluorescence intensity in IH group was obviously higher than that in G and C groups (P < 0.01). (2) Mitochondrial membrane potential: the fluorescence intensity in IH group was 32 +/- 7, which was obviously lower than that in G and C groups (52 +/- 4, 73 +/- 4, respectively, P < 0.01). (3) The condition of mPTP: the intensity in IH group was 27 +/- 4, which was obviously lower than that in G and C groups (62 +/- 8, 90 +/- 7, respectively, P < 0.01). (4) The activity of caspase-3: the activity of caspase-3 in IH group was obviously higher than that in G and C groups (P < 0.01). CONCLUSION: Glycine can inhibit apoptosis in cardiomyocytes subjected to ischemia and hypoxia,and the effect may be attributable to changes in mitochondrial membrane potential, lessening opening of mPTP, alleviation of calcium overload , and decrease in activity of caspase-3.

Impaired single cell firing and long-term potentiation parallels memory impairment following recurrent seizures.

Patients with epilepsy are at substantial risk for memory impairment. Animal studies have paralleled these clinical observations, demonstrating impaired hippocampal function as measured by spatial memory in rodents subjected to seizures. However, the mechanism of seizure-induced hippocampal impairment is unclear. Here we investigated the effects of recurrent seizures on water-maze performance, a behavioural measure of learning and memory, long-term potentiation (LTP; considered a test of synaptic plasticity and memory) and place-cell firing patterns, a single-cell indicator of spatial memory. LTP and CA1 place-cell activity were examined in separate groups of freely moving rats, before and after 10 flurothyl-induced seizures. Water maze performance was examined in a third group of rats, five with previously induced seizures and five controls. Recurrent flurothyl seizures were associated with marked impairment in LTP and a reduction in the frequency of the peak theta power. Compared to baseline recordings, place-cell firing patterns following recurrent seizures were significantly less precise, had lower firing rates and were less stable. Impaired place-cell firing was seen as early as after two seizures and persisted at least 72 h after the last seizure. Water-maze performance was also significantly impaired in animals that underwent recurrent seizures. No cell loss or synaptic reorganization was observed in the hippocampus or in several other cortical areas that are vulnerable to seizures. These results demonstrate that relatively brief excitatory events, not producing visible cell damage, can nevertheless cause long-lasting changes in hippocampal physiology, observable as impairments in place-cell function, LTP and spatial memory.

Postictal single-cell firing patterns in the hippocampus.

PURPOSE: Patients with epilepsy have varying degrees of postictal impairment including confusion and amnesia. This impairment adds substantially to the disease burden of epilepsy. However, the mechanism responsible for postictal cognitive impairment is unclear. The purpose of this study was to study single-cell firing patterns in hippocampal cells after spontaneous seizures in rats previously subjected to status epilepticus. METHODS: In this study, we monitored place cells and interneurons in the CA1 region of the hippocampus before and after spontaneous seizures in six epileptic rats with a history of status epilepticus. Place cells fire action potentials when the animal is in a specific location in space, the so-called place field. Place cell function correlates well with performance in tasks of visual-spatial memory and appears to be an excellent surrogate measure of spatial memory. RESULTS: Twelve spontaneous seizures were recorded. After the seizures, a marked decrease in firing rate of action potentials from place cells was noted, whereas interneuron firing was unchanged. In addition, when place cell firing fields persisted or returned, they had aberrant firing fields with reduced coherence and information content. In addition to postictal suppression of firing patterns, seizures led to the emergence of firing fields in previously silent cells, demonstrating a postictal remapping of the hippocampus. CONCLUSIONS: These findings demonstrate that postictal alterations in behavior are not due solely to reduced neuronal firing. Rather, the postictal period is characterized by robust and dynamic changes in cell-firing patterns resulting in remapping of the hippocampal map.

Effect of interictal spikes on single-cell firing patterns in the hippocampus.

PURPOSE: The interictal EEG spike(s) is the hallmark of the epileptic EEG. While focal interictal spike (IS) have been associated with transitory cognitive impairment, with the type of deficit dependent on where in the cortex the IS arises, the mechanism by which IS result in transitory dysfunction is not known. The purpose of this study was to determine the effect of IS on single-cell firing patterns in freely moving rats with a prior history of seizures. METHODS: We studied IS in two seizure models; pilocarpine-induced status epilepticus and recurrent flurothyl models. The effect of spontaneous hippocampal spikes on action potentials (APs) of CA1 cells in rats walking in a familiar environment was investigated using 32 extracellular electrodes. We also compared the effect of spikes on two types of hippcampal cells; place cells that discharge rapidly only when the rat's head is in a specific part of the environment, the so-called firing field, and interneurons, which are a main source of inhibition in the hippocampus. RESULTS: IS were associated with a decreased likelihood of AP compared with IS-free portions of the record. Compared to pre-IS baseline, IS were followed by significant decreases in CA1 APs for periods up to 2 s following the IS in both models. When occurring in flurries, IS were associated with a pronounced decrease in APs. The response to IS was cell-dependent; IS resulted in decreases in AP firing after the IS in interneurons but not place cells. CONCLUSIONS: This study demonstrates that IS have substantial effects on cellular firing in the hippocampus and that these effects last far longer than the spike and slow wave. Furthermore, the effect of IS on cellular firing was cell specific, affecting interneurons more than place cells. These findings suggest that IS may contribute to seizure-induced cognitive impairment by altering AP firing in a cell-specific manner.

Effect of levetiracetam on visual-spatial memory following status epilepticus.

Status epilepticus (SE) is often followed by severe cognitive impairment, including memory impairment. Previous studies have shown that SE is associated with impairment of single cells in the hippocampus that fire action potentials when the animal is in a specific location in space, the so-called place cells, and that place cell function correlates well with performance in tasks of visual-spatial memory. Place cell patterns therefore appear to be an excellent measure of spatial memory and may serve as a tool to assess seizure-induced impairment in memory. In this study we determined the relationship between visual-spatial memory and place cell function following SE. In addition, we determined if levetiracetam (LEV), an antiepileptic drug with a novel mechanism of action, can improve cognitive function and place cell firing patterns when administered following SE. SE was induced in adult male rats which were then randomized to post-SE treatment with LEV or normal saline (NS) treatment for 14 days. Non-SE control rats also were randomized to LEV or NS. Following discontinuation of LEV rats were tested for visual-spatial memory in the Morris water-maze and then underwent unit recording in the CA1 region of the hippocampus. Brains were then evaluated for cell loss and mossy fiber sprouting. SE was associated with severely impaired performance in the water-maze with SE rats demonstrating no learning over four days of testing. Paralleling this memory deficit was a marked disturbance in firing patterns of pyramidal neurons in CA1. Non-SE rats learned quickly over four days of water-maze testing and had normal pyramidal cell firing patterns. LEV had no major effects on water-maze performance or place cell function. Histopathological examination of the brains showed severe cell loss in CA1 in all of the SE rats with lesser degrees of injury in CA3 and the hilus. LEV treatment resulted in less histological damage in the hippocampus but had no effect on visual-spatial function or place cell physiology in either control or SE rats.

[Killing effect of PNP/MeP-dR suicide gene system driven by an AFP promoter AF0.3 on AFP-positive hepatoma cells].

BACKGROUND & OBJECTIVE: Alpha-fetoprotein (AFP) promoter-driven target gene could be specifically expressed in AFP-positive hepatoma. Escherichia coli purine nucleoside phosphorylase/6-methylpurine-2-deoxyriboside (PNP/MeP-dR) suicide gene system has powerful killing effects on tumor cells. This study was to investigate the specific killing effect of PNP/MeP-dR suicide gene system driven by an AFP promoter, AF0.3, on AFP-positive hepatoma cells. METHODS: Inserting PNP gene into pAF0.3, a eukaryotic expression vector containing PNP gene, pAF0.3/PNP, was constructed. Then it was transfected into AFP-positive HepG2 and AFP-negative SMMC7721 hepatoma cell lines, respectively. Two cell lines HepG2/AF0.3-PNP and SMMC7721/AF0.3-PNP, stably transfected with PNP gene, were obtained with G418 selection. The expression of PNP gene was detected by reverse transcription-polymerase chain reaction (RT-PCR). The proliferation of HepG2/AF0.3-PNP and SMMC7721/AF0.3-PNP cells was determined with trypan blue exclusion. The sensitivity of the cells to MeP-dR and the bystander effects were assessed with MTT assay and flow cytometry (FCM). The enzymatic activities of PNP gene products were determined with high performance liquid chromatography (HPLC). RESULTS: Whether hypoxia or normoxia, HepG2/AF0.3-PNP cells were sensitive to MeP-dR, whereas SMMC7721/AF0.3-PNP cells were not. Under both conditions, obvious cytotoxic effects on HepG2 cells were observed when the proportion of HepG2/AF0.3-PNP cells in the mixture reached 25%. But there were no similar effects on SMMC7721 cells under the same conditions. HPLC assay showed that the product of PNP gene driven by AF0.3 promoter could convert a spot of MeP-dR into 6-MP in HepG2 cells, but not in SMMC7721 cells. CONCLUSION: PNP/MeP-dR system, driven by AF0.3 promoter, has powerful killing effect on AFP-positive hepatoma HepG2 cells.