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OBJECTIVE: To investigate the effect of acupuncture stimulation of head acupoints "Jin San Zhen" (JIN's Three Acupuncture Needles Therapy) on behavior reactions, hippocampal neuronal autophagy and expression of autophagy associated proteins (Beclin-1 and light chain 3 Ⅱ/Ⅰ [LC 3 Ⅱ/Ⅰ]) in rats with hypoxic-ischemic brain damage (HIBD) due to fetal intrauterine distress, so as to reveal its underlying mechanisms in improving neonatal HIBD. METHODS: Pregnant SD rats were used in the present study. The HIBD model was established by delayed caesarean delivery and bilateral uterine arteries clipping for 10 minutes. The HIBD rats were randomly divided into model group and acupuncture groups (n＝9 rats in each group). The other 9 rats delivered naturally were used as the normal control group. On day 14 after delivery, the neonatal rats in the acupuncture group received acupuncture stimulation of head acupoints ("Nao San Zhen""Nie San Zhen" and "Zhi San Zhen") by twirling each needle leftward and rightward for 10 times, once a day for 14 d. The open field test and Morris water maze test were used to determine the locomotive activity and spatial learning-memory ability, respectively. The ultrastructure and autophagosomes in the hippocampal neurons were observed by transmission electron microscope. The contents and expression levels of Beclin-1 and LC3 Ⅱ/Ⅰ in the hippocampus tissues were detected by flow cytometry and Western blot, separately. RESULTS: Compared with the normal control group, the time to go out of the central region of open field test, and the escape latency and duration of first platform-quadrant-crossing of spatial exploration of Morris water maze tests were significantly increased (P<0.01，P<0.05，P<0.001), and the total distance and number of activities in the central region, and the target quadrant resistance time and number of platform-cros-sing remarkably decreased in the model group (P<0.01, P<0.05), suggesting a decline of both locomotor activity and learning-memory ability after modeling. The expression level (%) of Beclin-1 protein and ratio of LC3 Ⅱ/Ⅰ proteins were considerably increased in the model group (P<0.01). Following acupuncture interventions, the locomotor activity and spatial learning-memory ability were obviously increased (P<0.05，P<0.01，P<0.001), and the expression of Beclin-1 protein and ratio of LC3 Ⅱ/Ⅰ were further up-regulated relevant to the model group (P<0.001). Moreover, ultrastructural observation showed serrated change of nuclear membrane and widened perinuclear space, vacuolization in the mitochondria, dilation of endoplasmic reticulum and increase of autophagosomes in the hippocampal neurons in the model group. These situations were relatively milder in the acupuncture group. CONCLUSION: Acupuncture of head acupoints of "JIN San Zhen" may increase locomotor activity and learning-memory abi-lity in rats with HIBD due to fetal intrauterine anoxia, which is closely with its effect in promoting hippocampal neuronal autophagy via up-regulating the expression of Beclin-1 and LC3 Ⅱ/Ⅰ.
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Objective To explore the perinatal risk factors of early onset thrombocytopenia (EOT) in full-term small for gestational age infants. Methods A 1:1 or 1:2 matched case control study was carried out. A total of 93 full-term small for gestational age infants with EOT were selected from April 2008 to July 2014 as the case group, and the non EOT full-term small for gestational age infants with the birth weight difference <250 g and the gestational age difference <3 days were selected as the control group. The clinical data during perinatal period and laboratory examination results after admission were collected retrospectively. And the differences between the two groups were compared. Results The incidence of intrauterine distress (41.9％ vs. 25.8％, χ2=7.35, P=0.007), amniotic fluid contamination (39.8％ vs. 27％, χ2=4.66, P=0.031), and early-onset sepsis (39.8％ vs. 27％, χ2=4.66, P=0.031) were significantly higher in the case group than those in the control group. Conditional logistics regression analysis showed that intrauterine distress (β=0.60, OR=1.82, 95％CI=1.04～3.17, P=0.035) and early-onset sepsis (β=1.69, OR=5.44, 95％CI=1.11～26.76, P=0.037) were related to EOT. Conclusions Intrauterine distress and early-onset sepsis are risk factors for the onset of EOT in full-term small for gestational age infants.
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Aim To observe the neuroprotective effect of different doses of propofol on ischemic fetal rat brain.Methods Eighteen healthy pregnant SD rats were randomly allocated into the following six groups with three rats in each.Group S: sham operation group, Group IR: ischemia/reperfusion group, Group P1~P3: different doses of propofol groups, Group B: bicuculline group.In group S and group IR, 1 ml saline solution was administered via caudal vein.In group P1~P3, 10, 30, 50 mg·kg-1 of propofol was administered via caudal vein respectively.In group B, when 50 mg·kg-1 propfol was administered via caudal vein, 5 mg·kg-1 bicuculline was injected intraperitoneally at the same time.Bilateral uterine ovarian arteries were clamped for 11 mins to make intrauterine distress model of the fetal rats.The brains of fetal rats were removed after 3 days of reperfusion.Brain sections(5 μm thick) were mounted and stained with Hematoxylin and eosin(HE).The profile of the hippocampus CA1 was evaluated under a light microscope and neuronal Lesion-index(LI) was calculated.MDA content of fetal rat brain was detected by thiobarbituric acid reaction method to determine the lipid peroxidation degree of brain.Results LI was (7.2±0.9) and MDA was (3.86±0.20) μmol·g-1 in group S.LI was 71.9±2.8 and the content of MDA was (9.10±0.45) μmol·g-1 in group IR, which increased significantly compared with those in group S(P<0.01).LI was (40.8±2.6), (21.4±1.4), (20.1±1.3) and the content of MDA was (7.32±0.41), (5.65±0.27), (5.44±0.28) μmol·g-1 in propofol groups, which decreased significantly compared with those in group IR(P<0.05).LI and the content of MDA was (51.2±2.3), (7.54±0.31) μmol·g-1 in group B,respectively, reversing partly the neuroprotevtive effect of propofl.Conclusion Propofol could protect the neurons in hippocampus CA1 region of fetal rat against intrauterine distress by reducing the concentration of MDA in the brain.
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Objective To investigate the effects of penehyclidine hydrochloride (PHCD)on cerebral ischemia-reperfusion injury induced by intrauterine distress in fetal rats.Methods Eighty mature fetal rats weighing 4.52-4.81 g were randomly divided into four groups (n =20):sham opera-tion group(group S),PHCD control group (group S+ P),cerebral IR group (group IR),PHCD treatment group(group IR+P).Fetal rat intrauterine distress model was set up by clamping bilateral uterine horn vessels of pregnant rats.PHCD 2 mg/kg was injected in pregnant rat’s gluteus at 30 min before intrauterine distress model was set up in group IR+P,the same volume saline was injected in pregnant rat’s gluteus before shame operation in group S,the same volume PHCD was injected in pregnant rat’s gluteus before shame operation in group S+P.Fetal rats were decapitated at 12 h after the reperfusion,the peripheral blood of fetal rats was detected by blood gas analysis (including PH, PaO 2 ,PaCO 2 ,Lac);the infarct volume and the infarct volume fraction were detected by TTC stai-ning;pathological changes in lung tissue were observed by HE staining;the TNF-α,IL-6 content in the brain were detected by ELISA;the expression of NF-κB mRNA was detected by quantitative Real-time PCR,the expression of NF-κB p65 protein was detected by Western-blotting.Results The blood PH,PaO 2 in group IR and IR+P were lower than group S and S+P,the blood PH,PaO 2 in group IR+P was higher than group IR.Compared with group S and group S+P,the blood PaCO 2 , Lac,the infarct volume and the infarct volume fraction,the concentration of TNF-αand IL-6,the ex-pression of NF-κB mRNA and protein were significantly increased in group IR and IR+P (P <0.05), and those in group IR+P were lower than group IR (P <0.05 ).The pathological changes in brain tissue were significantly attenuated in group IR + P (P < 0.05 ).Conclusion Pretreatment with PHCDcouldattenuatecerebralischemia-reperfusioninjuryoffetalratsinducedbyintrauterinedistress. ThemechanismscouldrelatetotheinhibitionofNF-κBsignalingpathwayinbraintissues.
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Objective:To evaluae the unification of endoplasmic reticulum and mitochondrium in fetal rat cerebral neuron apoptosis induced by intrauterine distress. Methods:Fetal rat intrauterine distress model was established. The samples were randomly divided into normal group and ischemia-reperfusion group(IR). Expression of GRP78,caspase-9,caspase-12,and cytoron C protein were detected by western blot technique. Results:The expression of GRP78 on corresponding time in IR group was significantly higher than that in normal group. And it reached the maximum at 3h of reperfusion. The expression of GRP78 reduced step by step as reperfusion time extended. The expression of caspase-9 and caspase-12 at corresponding time in IR group were significantly higher than those in normal group. Their expression increased rapidly and reached the maximum at 12h of reperfusion. The expression of cytoron C in kytoplasm at corresponding time in IR group was significantly higher than that in normal group and reached the maximum at 12h of reperfusion. Conclusion:The synergistic effect of endoplasmic reticulum and mitochondrium plays an important role in fetal rat cerebral neuron apoptosis induced by intrauterine distress.
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Objective:To investigate the role of glucose-regulated protein 78(GRP78) and caspase-9 and-12 in intrauterine distress-induced hypoxic-ischemic cerebral damage in fetal rats.Methods:The fetal rat model of intrauterine distress was constructed and the fetal rats were randomly divided into a normal,a sham operation and an ischemia-reperfusion(IR) group.Neuron apoptosis was analyzed by in situ end-labeled DNA(TUNEL).The expressions of caspase-9 and-12 and GRP78 proteins in the hippocampus CA1 area were detected by immunohistochemical staining.Results:Ischemia-reperfusion damage induced classic neuron apoptosis and the number of the apoptotic neurons in the hippocampus CA1 area increased with the progression of reperfusion.The expressions of GRP78 and caspase-9 and-12 were weak in the normal and sham operation group.In the IR group,the expression of GRP78 reached the peak value 3 hours after the reperfusion and then decreased gradually;the intensity of caspase-12 was increased rapidly while that of caspase-9 elevated very little within 3 hours,but both reached the peak value at 12 hours.Conclusion:Intrauterine distress-induced hypoxic-ischemic cerebral damage in fetal rats may trigger the homeostatic control system in the endoplasmic reticulum through the increased expression of GRP78.The apoptotic pathway mediated by caspase-12 in the endoplasmic reticulum may be one of the mechanisms underlying cerebral ischemic injury.
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Objective:Evaluating the effect of nitric oxide synthase(NOS) inhibitors on fetal rat neuron apoptosis during intrauterine distress and childhood's learning and memory.Methods:① Set up fetal rat intrauterine distress model.②Divide into the control group,reperfusion group,treatment group [injected N omega-nitro-L-arginine(L-NNA) 4mg/kg into pregnant rat's abdomen before ischemia and injected aminoguanidine(AG) 500mg/kg before operation],long-term observation group Ⅰ?Ⅱ?Ⅲ(the part of rats in control group,reperfusion group,treatment group were performed cesarean section on pregnant 19 days and bred to 40 day of age after delivery).③Measuring the content of cytochrome c in cytosol and examined the ability of learning and memory function.Results:①The content of cytochrome c in reperfusion group(6/12/24h) and treatment group(6/12/24h) were higher than those in the control group(P0.05).(3) Crossing platform times in long-term observation groupⅡ were fewer than those in long-term observation group Ⅰ?Ⅲ(P 0.05).Conclusion:NOS inhibitors play a obvious protective role in long-term intelligence of fetal rat after delivery through relieved neuron apoptosis during hypoxic ischemia encephalopathy.NOS inhibitors can be a valuable therapy method in intrauterine distress treatment.