Objective Detection of S-100B protein in different degree meconium pollution level in serum of children with value.MethodsIn 2012 June to 2013 December in our department were simple meconium stained amniotic fluid in term newborns in 73 cases, and set up for the observation group, and according to the amniotic fluid pollution degree is divided into 47 cases of amniotic fluid of Ⅰ-Ⅱdegree pollution group and 26 cases in grade Ⅲmeconium group during the same period, selected 20 cases without amniotic fluid contamination in term healthy newborns for the control group, the groups were compared in S-100B protein content difference.ResultsⅢmeconium stained amniotic fluid were within 6h serum S-100B was significantly higher than that in control group(P<0.01), and the degree of contamination of amniotic fluidⅠ-Ⅱgroup compared with the control group, no significant difference(P>0.05).Ⅲ meconium stained amniotic fluid in children with 72h also increased.ConclusionThird degree meconium stained amniotic fluid but normal Apgar score of newborns may still exist in clinical brain injury, so pay close attention to.

In order to find the cardiotonic constituents of lateral roots of Aconitum carmichaelii Debx., the investigation was carried out. Silica gel column chromatography, Sephadex LH-20, medium-pressure MCI and reverse phase ODS column chromatography were used to separate the 90% EtOH extract of the lateral roots of Aconitum carmichaelii Debx. The structures of the isolated compounds have been identified by chemical properties and spectroscopic analyses. Ten compounds were isolated and their structures were elucidated as benzoic acid-5-hydroxy-2-benzoyl-amino methyl ester (1), honokiol (2), pinoresinol (3), salicylic acid (4), p-hydroxy-cinnamic acid (5), songorine (6), karakoline (7), mesaconitine (8), hypaconitine (9) and 14-benzoylhypaconitine (10), separetely. Compound 1 is a new compound and its structure has been established by NMR, HR-ESI-MS, UV, IR and X-Ray. Compound 2-5 are isolated from the lateral roots of Aconitum carmichaelii Debx. for the first time.
Aconitum ; chemistry ; Cardiotonic Agents ; chemistry ; isolation & purification ; Plant Roots ; chemistry

OBJECTIVETo explore the effect of ligustrazine on the migration of bone marrow mesenchymal stem cells (BMSCs) and protein expressions of matrix metalloproteinase-2 and-9 (MMP-2 and MMP-9) in vitro.

METHODSBMSCs were in vitro isolated and cultured using whole bone marrow adherent method, and phenotypes [surface positive antigens (CD29 and CD90) and negative antigens (CD34 and CD45)] identified using flow cytometry. BMSCs were divided into the blank control group, 25, 50, 100 µmol/L ligustrazine group, and the GM6001 group (100 µmol/L ligustrazine +MMPs inhibitor GM6001 ). The migration of BMSCs was tested by Transwell chamber test and wound healing assay after treated with ligustrazine for 24 h. The protein expressions of MMP-2 and MMP-9 were detected by Western blot.

RESULTSThe third passage BMSCs grew well in uniform morphology. The expression rate of CD29, CD90, CD34, and CD45 was 96.9%, 97.3%, 0.2%, and 3.0%, respectively. Compared with the blank control group, the number of migrated cells and relative distance of cell invasion increased, and the protein expressions of MMP-2 and MMP-9 were elevated in each ligustrazine group (P < 0.05, P < 0.01). Compared with 100 µmol/L ligustrazine group, the number of migrated cells and relative distance of cell invasion decreased in 25 and 50 µmol/L ligustrazine groups and the GM6001 group (P < 0.01). Protein expression of MMP-2 decreased in 25 and 50 µmol/L ligustrazine groups (P < 0.01).

CONCLUSIONLigustrazine could promote the migration of BMSCs in vitro, and its mechanism might be related to up-regulating expression levels of MMP-2 and MMP-9 protein.

Cell Movement ; Cells, Cultured ; Hematopoietic Stem Cells ; cytology ; drug effects ; Humans ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Pyrazines ; pharmacology ; Up-Regulation

OBJECTIVETo improve tooth pulp electrical stimulation rat model to study analgesic effect of drugs.

METHODSExpose lower mandible and incisor and isolate them with rubber dam, two holes with distance 1.5 mm were drilled below the cemento-enamel junction by a thin diamond bur in 22 adult male rats anesthetized with sodium pentobarbital. A pair of insulated stainless stimulating electrodes were inserted through the holes and knotted to fix them. Drilled holes were filled with zinc polycarboxylate cement. Finally, the stimulating electrodes were directed to the top of the skull. Pain threshold of rats was determined by an electrical stimulator under awake, free conditions to evaluate the stability of the model and the analgesic effect of drugs.

RESULTSA valid tooth pulp pain model were set up successfully in 22 rats. Rats regain consciousness at 1 to 2 hours after operation. The inserted electrodes keep efficiency for 6 weeks. Neither tissue morphology change nor inflammation cell infiltration can be found in tooth pulp after 6 weeks under the light microscope. Threshold voltages remained constant under the repeat electric stimulations within 300 min (P > 0.05, CV < 15%) until 4 weeks (P > 0.05, CV < 15%). The dose-dependent and time-dependent analgesic effects of capsaicin were showed in this model.

CONCLUSIONThe improved tooth pulp electrical stimulation rat model is easily performed and shows constant electrical stimulation-induced pain threshold and is suitable for research on analgesic effect of drugs.

Animals ; Dental Pulp ; Electric Stimulation ; Incisor ; Male ; Pain ; Rats

Melittin exhibits high antibacterial potency against drug-resistant bacteria. However, the clinical utility of melittin is limited by its serious hemolytic activity. Thus, the need for developing novel melittin analogues with high antimicrobial activity and low hemolytic activity has grown. We designed, synthesized, and evaluated 20 novel melittin analogues with varying hydrophobic, polar or positively charged amino acids. The results showed that 8 compounds had antimicrobial activity (MIC: 1-4 μg·mL^-1) against gram-positive pathogens equal to or better than that of melittin, and 16 compounds had low hemolytic activity (HC₅₀ ≥ 11.9 μg·mL^-1). Compounds 13 (MIC: 2-4 μg·mL^-1) and 15 (MIC: 1-2 μg·mL^-1) showed equal or better antimicrobial activity against both susceptible and resistant strains of Staphylococcus aureus and Enterococcus faecium compared to melittin (MIC: 4 μg·mL^-1). Compound 13 (HC₅₀: 24.0 ± 4.3 μg·mL^-1) displayed noticeably decreased hemolytic activity compared to melittin (HC₅₀: 5.3 ± 0.4 μg·mL^-1). This work established a base for further study on the structure-activity relationships and structure-toxicity relationships of melittin.

Aged ; Biopsy, Needle ; methods ; Coal Mining ; Humans ; Lung ; pathology ; Lung Neoplasms ; complications ; diagnosis ; Middle Aged ; Pneumoconiosis ; complications

OBJECTIVETo study the expression of nNOS and ultrastructural changes in the penile tissue of rats with prolactinoma-induced erectile dysfunction (ED).

METHODSWe established the model of prolactinoma in 20 male Westar rats by peritoneal injection of diethylstilbestrol (DES) and treated the control rats with normal saline (n = 10) or sterilized arachis oil (n = 10). After 8 weeks, we performed the apomorphine test and measured the weight of the pituitary gland and the levels of serum prolactin (PRL) and testosterone (T) to confirm the successful construction of the prolactinoma-induced ED model. Then we determined the expression of nNOS in the penile tissue by immunohistochemistry and examined the ultrastructural changes of the penile cavernosum under the transmission electron microscope.

RESULTSThe prolactinoma-induced ED model was successfully established in 15 rats. The weight of the pituitary gland was significantly increased in the rats treated with DES as compared with the normal saline and sterilized arachis oil controls ([46.7 ± 15.5] vs [11.7 ± 2.4] and [12.4 ± 2.3] mg, both P < 0.05). The level of serum PRL was markedly higher while that of T remarkably lower in the former than in the latter two groups ([1,744.9 ± 304.5] vs [11.5 ± 2.4] and [10.6 ± 1.9] ng/ml, both P < 0.0l; [1.54 ± 0.46] vs [3.11 ± 1.08] and [3.04 ± 1.11] ng/ml, both P < 0.05). The rate of penile erection was significantly reduced in the prolactinoma-induced ED model rats in comparison with the normal saline and arachis oil controls (16.7% vs 100% and 87.5%, both P < 0.05), and so was the expression of nNOS in the penile tissue (0.024 ± 0.011 vs 0.066 ± 0.019 and 0.058 ± 0.021, both P < 0.05). Transmission electron microscopy manifested significant ultrastructural changes in the endothelial and smooth muscle cells of the cavernous tissue in the prolactinoma-induced ED models.

CONCLUSIONThe ultrastructural changes of the penile cavernous tissue and the reduced expression of nNOS in penile tissue may be the most important mechanisms of prolactinoma-induced ED in rats.

Animals ; Apomorphine ; Carcinogens ; Diethylstilbestrol ; Erectile Dysfunction ; etiology ; Humans ; Male ; Myocytes, Smooth Muscle ; ultrastructure ; Nitric Oxide Synthase Type I ; metabolism ; Organ Size ; Penile Erection ; Penis ; enzymology ; ultrastructure ; Pituitary Neoplasms ; chemically induced ; complications ; Prolactin ; blood ; Prolactinoma ; chemically induced ; complications ; Rats ; Rats, Wistar ; Testosterone ; blood

OBJECTIVEThe application and therapeutic effect of hyperbaric oxygen (HBO) in hypoxic-ischemic brain damage (HIBD) remains controversial. Previous studies have focused on the early pathological and biochemical outcomes and there is a lack of long-term functional evaluation. This study was designed to evaluate the long-term pathological and behavioral changes of early HBO therapy on neonatal rats with HIBD.

METHODSPostnatal 7 days (PD7) rat pups were randomly assigned into Control (n=18), HIBD (n=17) and HBO treatment groups (n=17). HIBD was induced by ligating the left common carotid, followed by 2 hrs hypoxia exposure in the HIBD and HBO treatment groups. The Control group was sham-operated and was not subjected to hypoxia exposure. The HBO therapy with 2 atmosphere absolutes began 0.5-1 hr after HIBD in the HIBD treatment group, once daily for 2 days. The spatial learning and memory ability were evaluated by the Morris water maze test at PD37 to PD41. The morphological and histological changes of the brain, including brain weight, survival neurons, AchE positive unit and NOS positive neurons in hippocampal CA1 region, were detected at PD42.

RESULTSThe rats in the HIBD group displayed significant morphological and histological deficits, as well as severe spatial learning and memory disability. In the Morris water maze test, the mean escape latency were longer (56.35 +/- 22.37 s vs 23.07 +/- 16.28 s; P < 0.05) and the probe time and probe length were shorter in the HIBD group (29.29 +/- 6.06 s vs 51.21 +/- 4.59 s and 548 +/- 92 cm vs 989 +/- 101 cm; both P < 0.05) compared with the Control group. The left brain weight in the HIBD group was lighter than that in the Control group (0.601 +/- 0.59 g vs 0.984 +/- 0.18 g; P < 0.05). The survival neurons in the hippocampal CA1 region were less (100 +/- 27/mm vs 183 +/- 8/mm; P < 0.05), as well as the AchE-positive unit and NOS-positive neurons (18.50 +/- 2.24% vs 27.50 +/- 2.18% and 19.25 +/- 4.33 vs 33.75 +/- 5.57 respectively; P < 0.05) after HIBD. Early HBO treatment improved the abilities of spatial learning and alleviated the morphological and histological damage. The mean escape latency (39.17 +/- 21.20 s) was shortened, the probe time (36.84 +/- 4.36 s) and the probe length (686 +/- 76 cm) were longer, and the brain weight (0.768 +/- 0.85 g), the survival neurons (133 +/- 25/mm) and the AchE-positive unit (21.94 +/- 2.73%) increased significantly compared with those of the HIBD group (P < 0.05).

CONCLUSIONSEarly HBO treatment resulted in a protective effect against HIBD-induced long-term brain morphological and histological deficits and spatial learning and memory disability.

Acetylcholinesterase ; analysis ; Animals ; Brain ; pathology ; Escape Reaction ; Female ; Hippocampus ; enzymology ; pathology ; Hyperbaric Oxygenation ; Hypoxia-Ischemia, Brain ; enzymology ; pathology ; therapy ; Male ; Maze Learning ; Nitric Oxide Synthase ; analysis ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate possible mechanism of angiogenesis in brain tissue of neonatal rat hypoxic-ischemic encephalopathy (HIE).

METHODSForty seven-day old neonatal rats were randomly assigned to hypoxic-ischemic (Model group) or sham treatment (Sham group), each group had 20 rats. Five rats from each group were sacrificed on days 1, 3, 7 and 14 after hypoxia-ischemia. Paraffin sections of the brain were stained with anti-endothelial cell, anti-proliferating cell nuclear antigen (PCNA) or anti-vascular endothelial growth factor (VEGF) by using single or double immunohistochemistry. The brain capillary density index (BCDI), brain proliferating capillary density index (BPCDI) and the expression of VEGF were analyzed under the microscope. The expression of VEGF and hypoxia-inducible factor-1alpha (HIF-1alpha) mRNA in hypoxic-ischemic side of the brain was measured by RT-PCR.

RESULTSBCDI around infarct brain tissue in the model group began to rise on day 3 and remained higher than that of the sham group from day 3 to day 14 [day 3: (9.80 +/- 1.05)/HPF vs. (4.90 +/- 0.66)/HPF, P < 0.01;day 14: (13.29 +/- 3.90)/HPF vs. (6.08 +/- 1.50)/HPF, P < 0.01]. Occasional proliferating capillary was found in brain tissue of normal neonatal rats. The density of proliferating brain capillary on day 3 and day 7 of Model group [(0.54 +/- 0.15)/HPF vs. (0.90 +/- 0.25)/HPF] were significantly higher than those of Sham group [(0.12 +/- 0.05)/HPF vs. (0.13 +/- 0.07)/HPF, P < 0.01]. VEGF was mainly expressed in the cytoplasm of neurons, capillary endothelial cells and pial cells. Viable neurons and endothelial cells in the infarct areas also expressed VEGF. The expression of VEGF mRNA in hypoxic-ischemic brain tissue was significantly higher than that of normal control (P < 0.01) and temporally preceded angiogenesis. The expression of VEGF mRNA at 12 hours of HIE model was significantly higher than that of normal control (1.56 +/- 0.27 vs. 0.95 +/- 0.21, P < 0.05). It reached its peak on day 1 and day 3 (1.85 +/- 0.31 vs. 1.86 +/- 0.39), significantly higher than that of normal control (P < 0.01), and decreased by day 7 and day 14, without significant difference compared with normal control (P > 0.05). The expression of HIF-1alpha mRNA was also up-regulated after hypoxic-ischemic treatment. The expression of HIF-1alpha mRNA (1.07 +/- 0.21) was significantly higher than that of normal control (0.64 +/- 0.28, P = 0.048) at 3-hour of HIE model, reached its peak on day 1 (1.73 +/- 0.42, P < 0.01), remained at high expression level on day 3 (1.44 +/- 0.36, P < 0.05) and began to decline by day 7 and day 14 when it was not significantly different from normal control.

CONCLUSIONSAngiogenesis exists in the brain tissue of neonatal rat HIE model. Up-regulation of VEGF expression mediated by HIF-1 may play an important role in the process of angiogenesis.

Animals ; Animals, Newborn ; Brain ; blood supply ; Brain Diseases ; etiology ; genetics ; metabolism ; Disease Models, Animal ; Hypoxia-Inducible Factor 1, alpha Subunit ; Hypoxia-Ischemia, Brain ; complications ; Immunohistochemistry ; Neovascularization, Pathologic ; etiology ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction ; Transcription Factors ; analysis ; genetics ; Vascular Endothelial Growth Factor A ; analysis ; genetics

The effect of Rhizoma curcumae oil on the learning and memory in rats exposed to chronic hypoxia and the possible mechanisms were investigated. The rats were divided randomly into 5 groups (14 animals in each group): control, chronic hypoxia, chronic hypoxia with low (5 mg/kg body weight), middle (10 mg/kg body weight) and high (20 mg/kg body weight) concentrations of Rhizoma curcumae oil injection. The animals undergoing chronic hypoxia were exposed to hypoxia in a hypoxic chamber containing 10% O(2) and 5% CO(2) for 10 h/d, lasting 28 d. Morris water maze (MWM) test was used to obtain the scores of leaning and memory. The superoxide dismutase (SOD) activity and malonaldehyde (MDA) content were determined in the serum and hippocampus as well as [Ca(2+)](i) in the hippocampus. The expression of phosphorylated Ca(2+)/calmodulin-dependent protein kinase II (p-CaMKII) in the hippocampus was evaluated by using immunohistochemistry and Western blot. Compared with the control group, the chronic hypoxia group showed the following changes: (1) The escape latency to the hidden platform was remarkably prolonged (P<0.05); (2) The content of MDA and [Ca(2+)](i) were obviously higher, but the activity of SOD and the expression of p-CaMKII were significantly lower (P<0.05, P<0.01). Compared with the chronic hypoxia group, groups with Rhizoma curcumae oil injection had the following changes: (1) The escape latency to the hidden platform was remarkably shorter in 10, 20 mg/kg body weight groups (P<0.05); (2) The content of MDA and [Ca(2+)](i) were markedly decreased in 5, 10, 20 mg/kg body weight groups (P<0.05, P<0.01), but the activity of SOD in the serum and the expression of p-CaMKII were significantly higher in 10, 20 mg/kg body weight groups (P<0.05, P<0.01). The results showed that the capacity of learning and memory was degraded following chronic hypoxia. The decrease in MDA content and [Ca(2+)](i) and (or) the increase in SOD activity and p-CaMKII expression might participate in the enhancing effect on learning and memory induced by Rhizoma curcumae oil.
Animals ; Calcium ; metabolism ; Calcium-Calmodulin-Dependent Protein Kinase Type 2 ; metabolism ; Curcuma ; chemistry ; Hippocampus ; metabolism ; Hypoxia ; physiopathology ; Learning ; drug effects ; Malondialdehyde ; metabolism ; Memory ; drug effects ; Plant Oils ; pharmacology ; Rats ; Rhizome ; chemistry ; Superoxide Dismutase ; metabolism