To study in vitro anti-influenza viral activities of Chinese traditional patent medicines for influenza prevention and treatment, neuraminidase (NA) activity assay was used to examine NA inhibitory activity of 33 Chinese traditional patent medicines through fluorimetric assay, and influenza virus induced cytopathic effect (CPE) inhibition assay was used to verify their anti-influenza viral activities in vitro. The assay results showed that most liquid preparations displayed relatively high NA inhibitory activities, such as Shuanghuanglian oral liquid, Qingkailing oral liquid, Qingre Jiedu oral liquid, and Reduning injection. Among liquid preparations, Shuanghuanglian oral liquid not only displayed the highest NA inhibitory effect, but also exhibited obvious in vitro anti-viral activity in CPE experiment. Among solid preparations, Shuanghuanglian powder for injection showed the highest activity on NA inhibition, and Fufang Yuxingcao tablet showed relatively strong anti-influenza viral activity in CPE cells. From the results, it can be concluded that most Chinese traditional patent medicines possessed NA inhibitory activity, but only a few of them displayed significant in vitro anti-influenza viral activities. These results will provide important information for the isolation of active constituents, and for the clinical uses of Chinese traditional patent medicines for influenza treatment and prevention.

Bilirubin ; Child ; Duodenogastric Reflux ; diagnosis ; Esophageal Diseases ; diagnosis ; Esophagus ; metabolism ; Gastroesophageal Reflux ; diagnosis ; Humans

Adolescent ; Bilirubin ; metabolism ; Child ; Child, Preschool ; Duodenogastric Reflux ; metabolism ; Esophageal pH Monitoring ; Female ; Humans ; Male ; Stomach ; physiopathology

Objective To investigate the expression of pattern recognition receptor (PRR)of immune cells from the patients with enterovirus 71 ( EV 71 ) infection and the changes of cytokines. Methods Seventy-one patients and 20 age-matched healthy children were enrolled in the study. The patients were divided into three groups according to the critical degree: 20 mild patients, 25 severe patients and 26 critical patients. Real-time PCR were used to evaluate the levels of retinoic acidinducible gene Ⅰ ( RIG- Ⅰ ), melanoma differentitation-associated gene 5 ( MDA5 ) and cytokines IL-12, IFN-α mRNA expression in peripheral blood mononuclear cells (PBMC). The proportions of Toll like receptors(TLRs) on monocytes/macrophages (MC) , myloiddentritic cells (mDC) and plasmacytoiddentritic cells (pDC) were analyzed by flow cytometry. The concentrations of plasma cytokines( IL-12, IFN-α) were measured by ELISA. Results ( 1 ) The proportion of TLR7 is the unique TLR which is increased in mild patients and it was significantly elevated in MC, mDC and pDC in severe group (P＜0.05), TLR2, TLR3 and TLR4 also had an enhanced expression in MC and mDC. The enhanced expression of TLRs mentioned above had a decreased trend in critical patients. (2)Transcription levels of RIG- Ⅰ and MDA5 was significantly elevated in EV71 infected children.(3)The expression levels of DC-associated cytokines gene( IL-12 and IFN-α) have an increased trend in mild cases and these cytokines were remarkably increased in severe patients (P＜0.05), whereas decreased in critical cases (P＜0.05). ConclusionTLR7 might be the main PRR recognizing EV71 in immune cells and RIG-Ⅰ/MDA5 might also take part in the recognition of EV71. The exogenous or endogenous ligands released from bacterial infection and cell destruction could result in the activation of TLR2 or TLR4, inducing the inflammatory response.

Objective To study the change of CD4+ CD25+ Foxp3high regulatory T cells (Treg cells) and the molecules associated with Treg cells in different immune status in infant with sepsis, and to further clarity the pathogenesis of disturbed immune function in infant with sepsis. Method Totally 36 sepsis infants admitted in In-tensive Care Unit of Shenzhen Children' s Hospital from May 2007 to November 2007 and 16 age-matched healthy infants were collected for prospective study, after excluding autoimmune disease, immunodeficiency, inherited metabolic disorders, tumor, and drug-treatment that could affect immune function during lately 6 months. The study was approved by Ethics Committee of Shenzhen Children's Hospital. The 36 infants with sepsis were divided into two groups according to expression levels of HLA-DR in CD14-positive cells: DR-H group was defined as patients with HLA-DR > 30%, while DR-L group was defined as patients with HLA-DR < 30%. Expression levels of HLA-DR in CD14-positive cells and the proportion of Treg cells were analyzed by flow cytometry. Real-time PCR were used to evaluate the mRNA levels of Foxp3, CTLA-4,GITR, and IL-10 in CD4-posidve ceils. Statistical analysis was performed by one-way Anova. There was statistical difference with P < 0.05. Results The proportion of Treg cells in DR-L group was found to be significantly higher than that in healthy control or DR-H group (P <0.05).Compared with healthy control group or DR-H group, transcriptional levels of Foxp3, CTLA-4 and IL-10were significantly increased in DR-L group (P <0.05). The levels of GITR mRNA in DR-L group were detected to be higher than those in DR-H group (P < 0.05). Conclusions Aberrant increased proportion of Treg cells may be associated with suppressed immune status in infant with sepsis.

Objective To investigate the changes of CD4~+ T cell subset in the role of immuno-pathogenesis of type 1 diabetes mellitus(T1DM). Methods Real-time PCR was used to evaluate the ex-pression levels of transcriptional factors (T-bet, GATA-3, Foxp3, ROR-γt), cytokines (IFN-γ, IL-4, IL-10, IL-17A) and negative regulatory factors (CTLA-4, GITR) mRNA from CD4~+ T cells. The proportions of Th1, Th2, Tr and Th17 cells in peripheral blood were detected by flow cytometric analysis. Plasma cyto-kine (IFN-γ, IL-4, TGF-β and IL-6) concentrations were measured by enzyme-linked immunosorbent assay (ELISA). Results (1) The proportions of Th1 cells in peripheral blood from children with T1DM were siguificanfly increased than that of healthy controls, and proportions of Th2 were decreased (P < 0.01). There were no significant differences between diabetic patients and healthy controls regarding the proportions of Tr cells and Th17 cells(P >0.05). (2) Transcription levels of T-bet and IFN-γ mRNA were significantly up-regulated, while GATA3 and IL-4 were significantly down-regulated in children with T1DM. The mRNA expression levels of Tr negativity regulatory factors such as IL-10, CTLA-4 and GITR were lower in CD4~+ T cells from children with TIDM compared with the controls(P <0.01). There were no statistically differences to be observed in mRNA expression levels of ROR-γt and IL-17A genes between two groups(P > 0.05).(3) In comparison with controls, serum concentrations of IFN-γ or IL-4 were remarkable increased or de-creased respectively (P < 0. 01), while TGF-β and IL-6 did not change in children with T1DM (P > 0.05). Conclusion The Th1/Th2 imbalance might be play an important role in immunopathogenesis of T1DM. Functional deficiency of Tr cell might further exacerbate Th1/Th2 imbalance and lead to disturbance of cellu-lar immune response.

Objective To further explore the pathogenesis of disturbed adaptive immune response in infants with sepsis. Methods Forty-eight infants with sepsis and 26 age-matched healthy infants were enrolled in this study. The HLA-DR expression of CD14~+ monocyte, the proportion of CD4~+ CD25~+ Foxp3~(high) Tr cells and the proportion of Thl7 cells were measured by flow cytometry. Cytokines (IL-1β, IL-6, TNF-α, IL-10, TGF-β and IL-17A) were measured by ELISA. Real-time PCR were used to evaluate the mRNA levels of Foxp3, ROR-γt in CD4-positive cells and IL-17A. Forty-eight infants with sepsis were divided into two groups according to HLA-DR expression of CD14~+ monocyte: DR-H group ( ＞ 30% ) and DR-L group ( ＜ 30% ). Results The ratio of IL-10/TNF-α in DR-L group was higher than that in healthy control or DR-H group(P ＜0.05). The proportion of CD4~+ CD25~+ Foxp3~(high) Tr cells and mRNA expression of transcription factor Foxp3 in DR-L group was found to be significantly higher than that in healthy control or DR-H group(P＜0.05). The proportion of Thl7 cells, Serum concentration of IL-17A, the mRNA expression of IL-17A and transcription factor ROR-γt were significantly increased in DR-H group and DR-L group (P ＜ 0.05) , while there is no significant difference between DR-H and DR-L group( P ＞0.05). Serum levels of Th17-inducing cytokine such as IL-1β, IL-6 were significantly elevated in DR-H group and DR-L group (P＜0.05), while there is no significant difference between DR-H and DR-L group( P＞0.05). Serum level of CD4~+ CD25~+ Foxp3~(high) Tr-inducing cytokine TGF-p in DR-L group was higher than that in DR-H or healthy control group(P＜0. 05). Conclusion Over-activation of Th17 cells may be one of the factors causing aberrant increase of pro-inflammatory cytokine/chemotatic factor in infant with sepsis. The imbalance of CD4~+ CD25~+ Foxp3~(high) Tr cells/Th17 cells may be contributed to the pathogenic mechanism of mixed antagonist response syndrome ( MARS) in infant with sepsis. The changes of cytokine environment in infants with sepsis may be one of the factors causing the imbalance of CD4~+ CD25~+ Foxp3~(high) Tr cells/Th17 cells.

The purpose of the present work was to study hippocampal cellular electrophysiology involved in cortical epileptic networks reorganized by acute tetanization (60 Hz, 2 s, 0.4-0.6 mA) of the right posterior dorsal hippocampus (ATPDH). Experiments were performed on 35 adult male Sprague-Dawley rats. The ATPDH was used to establish epilepsy model. Four-channel recordings were done simultaneously, two channels for single unit recordings from bilateral hippocampi and other two channels for electrocorticogram (ECoG) or hippocampal electroencephalogram (EEG) recordings on the bilateral sides. The ATPDH bilaterally induced: (1) Epileptic ECoG activities that were originated from single unit afterdischarges of ipsilateral hippocampal neuron, followed by single unit afterdischarges of contralateral hippocampal neuron, finally produced seizure-like oscillations in the contralateral and ipsilateral ECoG. (2) 4-10 Hz rhythmic ECoG oscillations and 100-250 Hz hippocampal EEG oscillations, and asymmetric electrical activities of bilateral hippocampal neurons. (3) The hippocampal neuronal bursting that appeared its interspike interval (ISI) spot distribution in an irregular circular shape. The ipsilateral circular shape ISI distribution was more regular and occurred often, which was time-locked with the sinusoidal pattern of maximum peaks of hippocampal oscillatory trains. The results imply that the epileptic networks are reorganized bilaterally between ECoG and hippocarnpal EEG by the ATPDH. The hippocampal neuronal firing temporally encodes its information, in particular, the irregular circular shape ISI distribution during epileptic network reorganization.

The purpose of the present work was to study the effect of acute tetanization of the right caudate putamen nucleus (ATRC) on single neuronal interspike intervals (ISIs) in both laterodorsal thalamic nuclei (LDi), and electroencephalogram (EEG) wave interpeak intervals (IPIs) in both hippocampi (HPCi). Experiments were performed on 21 male Sprague-Dawley rats weighing 150~250 g. The seizures were induced by the ATRC (60 Hz, 2 s, 0.4~0.6 mA). Quadruple recordings were simultaneously carried out: two for single unit recordings from both LDi, and two for EEG recordings from both HPCi. The ATRC induced: (1) An interactive epileptic electrical network reconstructed in bilateral HPCi, which was driven by primary afterdischarges of single LD neuron. (2) A symmetric mirror-like ISI spot distribution of the LD neuronal firing before and after tetanus. (3) Gradually prolonged LD neuronal discharge intermittence was coherent with synchronous hippocampal EEG activities on the contralateral side. (4) Single LD neuronal spikes were phase- and time-locked to 20~25 Hz gamma oscillations in contralateral HPC. It suggests a particular temporal code patterning of single LD neuronal firing and its relationships to hippocampal EEG wave code in time series, the latter implies the LD neuronal encoding mechanisms of ATRC-induced epileptic electrical network in bilateral HPCi.
Action Potentials ; physiology ; Animals ; Caudate Nucleus ; physiology ; Electric Stimulation ; methods ; Electroencephalography ; Epilepsy ; etiology ; physiopathology ; Hippocampus ; physiology ; Lateral Thalamic Nuclei ; physiology ; Male ; Nerve Net ; physiology ; Neurons ; physiology ; Rats ; Rats, Sprague-Dawley ; Reaction Time ; physiology

The purpose of the present work was to study the role of unilateral hippocampal neural network in hippocampal epileptogenesis and its cellular mechanisms. Experiments were performed on 45 Sprague-Dawley adult rats. Acute tetanization (60 Hz, 2 s, 0.4 - 0.6 mA) of the right posterior dorsal hippocampus (ATPDH) was used to induce hippocampal epilepsy. The single unit discharges and the depth electrographs were synchronously recorded with a glass microelectrode and a pair of stainless concentric electrodes in the ipsilateral anterior dorsal hippocampus (HPC). The results demonstrated that: (1) some primary unit after-discharges were synchronized with electrographic after-discharges in the anterior dorsal HPC network after eight or nine tetanic trains were administered. Others desynchronized with 5 - 90 Hz primary depth electrographic after-discharges; (2) primary electrographic after-discharges were driven by primary unit after-discharges in the anterior dorsal HPC; (3) primary unit after-discharges were induced by brief primary electrographic after-discharges; and (4) plasticity of primary electrographic after-discharges and inhibition of single neuron firing were induced by repetitive ATPDH. The results suggest that hippocampal pathophysiologic network along the temporal-septal axis of the HPC is re-established by the repetitive ATPDH. There are plastic interactions between single neurons and its network during this re-establishment, which may be involved in the generation of "seizure oscillation". Over-activation of an intrinsic inhibition of the HPC along its temporal-septal axis might be involved in hippocampal network epileptogenesis.
Animals ; Electric Stimulation ; Epilepsy, Temporal Lobe ; physiopathology ; Evoked Potentials ; Hippocampus ; physiopathology ; Male ; Nerve Net ; physiopathology ; Neurons ; physiology ; Rats ; Rats, Sprague-Dawley