Bloodletting Puncture at Hand Twelve Jing-Well Points Relieves Brain Edema after Severe Traumatic Brain Injury in Rats via Inhibiting MAPK Signaling Pathway / 中国结合医学杂志

OBJECTIVE@#To investigate whether blood-brain barrier (BBB) served a key role in the edema-relief effect of bloodletting puncture at hand twelve Jing-well points (HTWP) in traumatic brain injury (TBI) and the potential molecular signaling pathways.@*METHODS@#Adult male Sprague-Dawley rats were assigned to the sham-operated (sham), TBI, and bloodletting puncture (bloodletting) groups (n=24 per group) using a randomized number table. The TBI model rats were induced by cortical contusion and then bloodletting puncture were performed at HTWP twice a day for 2 days. The neurological function and cerebral edema were evaluated by modified neurological severity score (mNSS), cerebral water content, magnetic resonance imaging and hematoxylin and eosin staining. Cerebral blood flow was measured by laser speckles. The protein levels of aquaporin 4 (AQP4), matrix metalloproteinases 9 (MMP9) and mitogen-activated protein kinase pathway (MAPK) signaling were detected by immunofluorescence staining and Western blot.@*RESULTS@#Compared with TBI group, bloodletting puncture improved neurological function at 24 and 48 h, alleviated cerebral edema at 48 h, and reduced the permeability of BBB induced by TBI (all P<0.05). The AQP4 and MMP9 which would disrupt the integrity of BBB were downregulated by bloodletting puncture (P<0.05 or P<0.01). In addition, the extracellular signal-regulated kinase (ERK) and p38 signaling pathways were inhibited by bloodletting puncture (P<0.05).@*CONCLUSIONS@#Bloodletting puncture at HTWP might play a significant role in protecting BBB through regulating the expressions of MMP9 and AQP4 as well as corresponding regulatory upstream ERK and p38 signaling pathways. Therefore, bloodletting puncture at HTWP may be a promising therapeutic strategy for TBI-induced cerebral edema.

Effects of Exendin-4 on the differentiation of neural stem cells from subventricular zone of adult mice in vitro / 中国应用生理学杂志

OBJECTIVE@#To study the effect of exendin-4(Ex-4) on the differentiation of neural stem cells(NSCs) in adult mouse subventricular zone(SVZ)and its mechanism .@*METHODS@#NSCs in the SVZ were derived from 5-week C57BL/6J mice and the expression of nestin was detected by immunofluorescence. The cell morphology was observed after the cells treatmed with 100 nmol/L Ex-4 for 14 days.The expressions of nestin and glucagon-like peptide-1 receptor (GLP-1R) were detected by immunofluorescence. GLP-1R was knocked down by using shRNA and the study was divided into four groups: control group, Ex-4 group, GLP-1R knockdown group, GLP-1R knockdown + Ex-4 group. After treatment with 100 nmol/L Ex-4 for 14 d, β-tublin III and glial fibrillary acidic protein (GFAP) were labeled by immunofluorescence and then the proportion of β-tublin III positive cells were counted. Western blot was used to detect the activation of cAMP-response element binding protein (CREB) in NSCs. In order to further study the effects of Ex-4 on mitogen-activated protein kinase(MAPK) and phosphatidylinositol 3-hydroxy kinase (PI3K) pathways, the cells were pretreated with MAPK inhibitor U0126 at a concentration of 0.07 μmol/L for 30 min or PI3K inhibitor LY294002 at 50 μmol for 2 h, respectively. The study was divided into six groups: control group, Ex-4 group, U0126 group, U0126 + Ex-4 group, LY294002 group, LY294002 + Ex-4 group. The activation of CREB in each group was detected by Western blot. The experiment was repeated three times independently.@*RESULTS@#NSCs were successfully extracted from SVZ of C57BL/6J mice. Immunofluorescence showed that nestin and GLP-1R were positive in NSCs. Compared with the control group, the proportion of neurons differentiated from Ex-4 group was higher. The percentage of neurons in GLP-1R knockdown + Ex-4 group was basically the same as that in control group (P＜0.01). The positive cells of beta-tublin III showed positive activation of GLP-1R and CREB. Western blot showed that CREB was significantly activated in the Ex-4 group, and knockdown of GLP-1R abolished its activation (P＜0.01). U0126 did not affect Ex-4-mediated CERB activation, and LY294002 significantly reduced Ex-4-mediated CREB activation (P＜0.01).@*CONCLUSION@#Ex-4 promotes the differentiation of NSCs into neurons in SVZ of adult mice through GLP-1R receptor, which may be achieved through PI3K/CREB pathway.

Comparative study on sepsis models induced by Escherichia coli subtypes / 天津医药

Objective To investigate the degrees of injury severity of sepsis models made by different kinds of Escherichia coli. Methods The 152 mice were randomly divided into control group, DH5α group, 44102 group, and 25922 group, with 38 rats in each group. DH5α group, 44102 group and 25922 group were intraperitoneally injected with 300 μL of Escherichia coli DH5α, 44102 and 25922 at the concentration of 1.0 × 109CFU/kg to prepare sepsis models of different kinds of Escherichia coli. Control group was injected intraperitoneally with the same amount of normal saline. (1) After 8 h, four mice were taken from each group for peripheral blood bacterial culture . (2) After 12 h, ten mice in each group were used for measuring serum levels of TNF-α and IL-6 by enzyme-linked immunosorbent assay (ELISA). (3) Western blot assay was used to determine the serum levels of high-mobility group protein (HMGB1) in four mice of each group. (4) Ten mice in each group were used to measure serum levels of alanine transaminase (ALT), aspartate aminotransferase (AST), creatinine (CR) and blood urea nitrogen (BUN) by automatic biochemical analyzer. (5) After liver, lung and kidney tissues were fixed with formaldehyde, hematoxylin-eosin (HE) staining was performed (n=10 for each group). Results In DH5α group, 44102 group and 25922 group, bacteria, inflammatory cytokines TNF-α, IL-6 and HMGB1 protein, liver and kidney indicators ALT, AST, CR and BUN showed a sequential increasing trend (P＜0.01). The severe degrees of alveolar structure damage, hepatic cell infiltration and renal glomerular atrophy were DH5α group, 44102 group and 25922 group in turn. There were no obvious damages of lung, liver or kidney tissues in control group. Conclusion Escherichia coli 25922 induces severe sepsis injury and can be used to study the animal models of the initial inflammatory phase of sepsis. Escherichia coli 44102 induces moderate damage of sepsis and can be used in animal models that do not require definitive sepsis staging experiments. Escherichia coli DH5α induces less damage of sepsis and can be used to explore immunosuppressive therapy of the animal model of sepsis.

Effects of ibuprofen on the growth and development of oligodendrocytes / 天津医药

Objective To study the effects of ibuprofen on the growth and development of oligodendrocytes. Methods A total of 6 clean and healthy adult female SD (Sprague Dawley) rats were used for extracting and culturing of oligodendrocytes(OLs).Lysophosphatidic acid(LPA)was then added,and the morphological changes of OLs pre-treatment and post-treatment were observed. Then 6 newborn rats (born 24-48 h) were used for mixed glial cell extraction from the cortex, then the OPCs were inoculated into the culture plates and randomly divided into control group, ibuprofen group, lysophosphatidic acid(LPA)group and LPA+ibuprofen group.After the adhering of the cells in each group for three days, cell morphology was observed,and the drugs were added as interventions.The control group was treated with normal saline, and the other 3 groups were added with saline solution of ibuprofen(100 μmol/L),LPA(1.0 μmol/L)and the mixture of them. The cell morphological changes were observed after 7-day intervention.The morphology of OPCs and OLs were observed by immunofluorescence staining through OPCs'specific immune markers (platelet-derived growth factor receptor alpha, PDGFR-α)and OLs'specific immune markers(myelin basic protein,MBP)along with cell count of mature OLs.Western blot assay was used to detect the relative expression level of MBP in each group. Results After the treatment with LPA to the mature OLs,protrusions were shrinking and became very sparse.The morphology of cells developed well in each group after cell adhering for 3 days. After drug intervention for 7 days, more cell protrusions and branches were observed in ibuprofen group and LPA+ibuprofen group than those of the control group and LPA group.The results of cell count showed that the number of MBP positive cells was significantly higher in the ibuprofen group and LPA+ibuprofen group than that in the control group and LPA group(P<0.01).The results of Western blot assay showed that the MBP protein expression was significantly less in LPA group than the other three groups (P<0.01), and the expression was significantly higher in the ibuprofen group than that of LPA+ibuprofen group (P<0.01). Conclusion LPA has a toxic effect on the growth and development of OPCs, and it has an inhibitory effect on the normal growth of mature OLs. A certain concentration of ibuprofen can significantly inhibit the cytotoxicity of LPA on OPCs and OLs,and promote the formation and maintenance of mature OLs.

Protective effect of oxiracetam on traumatic brain injury in rats / 中国应用生理学杂志

<p><b>OBJECTIVE</b>To study the role of oxiracetam on traumatic brain injury in rats.</p><p><b>METHODS</b>Thirty Wistar rats were randomly divided into 3 groups: sham operation group, model group and treatment group. Feeney method were used to establish traumatic brain injury (TBI) model in rats in model and treatment group, and rats in sham group were only broached without hydraumatic fitted. Rats in treatment group were successive administration for 21 days with oxiracetam (100 mg/kg, ig). Neurologic impairment scores were undertook after operation of 1 d, 4 d, 7 d, 14 d and 21 d, and Morris water maze test were proceeded during 15 to 19 days after operation. Average escape latency, searching time in target quadrant and number of crossing target platform in rats were recorded.</p><p><b>RESULTS</b>Neurologic impairment scores of rats in treatment group were significantly less than those of model group after operation of 7, 14 and 21 d (P < 0.05). Average escape latency of model group were significantly higher than those of sham operation group and treatment group (P < 0.05, P < 0.01). Searching time in target quadrant and number of crossing target platform of model group were lower than those of sham operation and treatment group (P < 0.05)).</p><p><b>CONCLUSION</b>Oxiracetam could decrease neural injury and increase ability of learning, memory and space cognition in traumatic brain injury rats.</p>

Role of stanniocalcin 1 in brain injury of coal-burning-borne fluorosis rats / 中国地方病学杂志

Objective To observe the change of stanniocalcin 1 (STC1) and calcium content in brain of coal-burning-borne fluorosis rats,and to explore the role of STC1 in brain injury of coal-burning-borne fluorosis.Methods Twenty four male SD rats were randomly divided into control,low,medium,and high fluoride groups according to body mass.Control group was fed conventional rat chow(fluorinated 1.3 mg/kg),and low,medium and high fluoride groups fed with fluorinated feed(20.0,40.0,60.0 mg/kg).All rats were given distilled water and feed ad libitum.One hundred and eighty days after modeling,STC1 protein and gene expression in the brain tissue of rats were detected using immunohistochemistry and RT-PCR and calcium content of brain tissue was detected.Results The cell positive rates of STC1 in low,medium,high fluoride groups [(48.10 + 2.11)％,(54.90 ± 1.73)％,(79.30 ± 3.71)％] were significantly higher than that of the control group[(24.70 + 3.53)％,all P ＜ 0.05],the cell positive rate of high fluoride group was significantly higher than that of the low and medium fluoride groups (all P ＜ 0.05).The STC1 mRNA expression of low,medium and high fluoride groups (0.58 ± 0.09,0.85 ± 0.17,1.75 ± 0.04) were significantly higher than that in the control group(0.37 ± 0.12,all P＜ 0.05),the STC1 mRNA expressions of high fluoride group was significantly higher than that of the low and medium fluoride groups (all P ＜ 0.05).The brain cortex calcium ion concentrations of low,medium and high fluoride groups[(138.62 + 4.19),(167.43 + 6.57),(189.45 + 3.72)nmol/L] were significantly higher than that in the control group [(101.47 + 9.46)nmol/L,all P ＜ 0.05],the brain cortex calcium ion concentrations of high fluoride group was significantly higher than that of the low and medium fluoride groups(all P ＜ 0.05),and the medium fluoride groups was higher than the low groups (P ＜ 0.05).Conclusion STC 1 may be involved in brain damage of coal-burning-borne fluorosis rats through regulating calcium balance.

The study in primary cultured astrocytes following fluid percussion injury / 中国应用生理学杂志

<p><b>OBJECTIVE</b>To investigate the morphology alterations and proteomics changes in primary astrocytes following fluid percussion injury.</p><p><b>METHODS</b>Primary cultures of astrocytes were prepared from cerebral hemispheres of 1-3 d-old SD rats, then, astrocytes were randomly divided into control group and injury group which were subjected to (0.2 +/- 0.01) MPa fluid percussion injury. The changes of protein expression pattern in astrocytes between injury and control groups were monitored with two dimensional gel electrophoresis.</p><p><b>RESULTS</b>Astrocytes' s abnonmalities of morphology after injury were apparent. The fluid percussion injury caused astrocytes edema, shrinkage, cell junction disconnection and necrosis at 2 h after injury. 24 h and 48 h after injury, most part of astrocytes's dendrites and soma became hypertrophy and showed a higher rate of cell proliferation. The dynamic proteomics changes were identified and total different 13 spots were detected in this study from the 2DE gels. The different displayed 5 spots were identified via MALDI-TOF: cofilin 1, destrin, phosphoglycerate mutase 1, NADH dehydrogenase (ubiquinone) 1 alpha subcomplex 10, annexin 1.</p><p><b>CONCLUSION</b>The obvious alteration of morphology and protein expression pattern in primary cultured astrocytes could be induced after fluid percussion injury. The differential proteins detected were probably related to stress responses.</p>