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This study aims to investigate the effect of Bombyx Batryticatus extract(BBE) on behaviors of rats with global cerebral ischemia reperfusion(I/R) and the underlying mechanism. The automatic coagulometer was used to detect the four indices of human plasma coagulation after BBE intervention for quality control of the extract. Sixty 4-week-old male SD rats were randomized into sham operation group(equivalent volume of normal saline, ip), model group(equivalent volume of normal saline, ip), positive drug group(900 IU·kg~(-1) heparin, ip), and low-, medium-, and high-dose BBE groups(0.45, 0.9, and 1.8 mg·g~(-1)·d~(-1) BBE, ip). Except the sham operation group, rats were subjected to bilateral common carotid artery occlusion followed by reperfusion(BCCAO/R) to induce I/R. The administration lasted 7 days for all the groups. The behaviors of rats were examined by beam balance test(BBT). Morphological changes of brain tissue were observed based on hematoxylin-eosin(HE) staining. Immunofluorescence method was used to detect common leukocyte antigen(CD45), leukocyte differentiation antigen(CD11b), and arginase-1(Arg-1) in cerebral cortex(CC). The protein expression of interleukin-1β(IL-1β), interleukin-4(IL-4), interleukin-6(IL-6), and interleukin-10(IL-10) was detected by enzyme-linked immunosorbent assay(ELISA). The non-targeted metabonomics was employed to detect the levels of metabolites in plasma and CC of rats after BBE intervention. The results of quality control showed that the BBE prolonged the activated partial thromboplastin time(APTT), prothrombin time(PT), and thrombin time(TT) of human plasma, which was similar to the anticoagulation effect of BBE obtained previously. The results of behavioral test showed that the BBT score of the model group increased compared with that of the sham operation group. Compared with the model group, BBE reduced the BBT score. As for the histomorphological examination, compared with the sham operation group, the model group showed morphological changes of a lot of nerve cells in CC. The nerve cells with abnormal morphology in CC decreased after the intervention of BBE compared with those in the model group. Compared with the sham operation group, the model group had high average fluorescence intensity of CD45 and CD11b in the CC. The average fluorescence intensity of CD11b decreased and the average fluorescence intensity of Arg-1 increased in CC in the low-dose BBE group compared with those in the model group. The average fluorescence intensity of CD45 and CD11b decreased and the average fluorescence intensity of Arg-1 increased in medium-and high-dose BBE groups compared with those in the model group. The expression of IL-1β and IL-6 was higher and the expression of IL-4 and IL-10 was lower in the model group than in the sham operation group. The expression of IL-1β and IL-6 was lower and the expression of IL-4 and IL-10 was higher in the low-dose, medium-dose, and high-dose BBE groups than in the model group. The results of non-targeted metabonomics showed that 809 metabolites of BBE were identified, and 57 new metabolites in rat plasma and 45 new metabolites in rat CC were found. BBE with anticoagulant effect can improve the behaviors of I/R rats, and the mechanism is that it promotes the polarization of microglia to M2 type, enhances its anti-inflammatory and phagocytic functions, and thus alleviates the damage of nerve cells in CC.
Subject(s)
Humans , Rats , Male , Animals , Interleukin-10 , Rats, Sprague-Dawley , Interleukin-4/metabolism , Bombyx , Interleukin-6/metabolism , Microglia/metabolism , Saline Solution/metabolism , Reperfusion Injury/metabolism , Brain Ischemia/metabolism , Cerebral Infarction , Reperfusion , NeuronsABSTRACT
When ischemia or hemorrhagic stroke occurs, astrocytes are activated by a variety of endogenous regulatory factors to become reactive astrocytes. Subsequently, reactive astrocytes proliferate, differentiate, and migrate around the lesion to form glial scar with the participation of microglia, neuron-glial antigen 2(NG2) glial cells, and extracellular matrix. The role of glial scars at different stages of stroke injury is different. At the middle and late stages of the injury, the secreted chondroitin sulfate proteoglycan and chondroitin sulfate are the main blockers of axon regeneration and nerve function recovery. Targeted regulation of glial scars is an important pathway for neurological rehabilitation after stroke. Chinese medicine has been verified to be effective in stroke rehabilitation in clinical practice, possibly because it has the functions of promoting blood resupply, anti-inflammation, anti-oxidative stress, inhibiting cell proliferation and differentiation, and benign intervention in glial scars. This study reviewed the pathological process and signaling mechanisms of glial scarring after stroke, as well as the intervention of traditional Chinese medicine upon glial scar, aiming to provide theoretical reference and research evidence for developing Chinese medicine against stroke in view of targeting glial scarring.
Subject(s)
Humans , Astrocytes , Axons/pathology , Cicatrix/pathology , Gliosis/pathology , Medicine, Chinese Traditional , Nerve Regeneration , Stroke/drug therapyABSTRACT
OBJECTIVE@#Our previous research showed that Naotaifang (a compound traditional Chinese herbal medicine) extract (NTE) has clinically beneficial effects on neurological improvement of patients with acute cerebral ischemia. In this study, we investigated whether NTE protected acute brain injury in rats and whether its effects on ferroptosis could be linked to the dysfunction of glutathione peroxidase 4 (GPX4) and iron metabolism.@*METHODS@#We established an acute brain injury model of middle cerebral artery occlusion (MCAO) in rats, in which we could observe the accumulation of iron in neurons, as detected by Perl's staining. Using assay kits, we measured expression levels of ferroptosis biomarkers, such as iron, glutathione (GSH), reactive oxygen species (ROS) and malonaldehyde (MDA); further the expression levels of transferrin receptor 1 (TFR1), divalent metal transporter 1 (DMT1), solute carrier family 7 member 11 (SLC7A11) and GPX4 were determined using immunohistochemical analysis, real-time quantitative polymerase chain reaction and Western blot assays.@*RESULTS@#We found that treatment with NTE reduced the expression levels of TFR1 and DMT1, reduced ROS, MDA and iron accumulation and reduced neurobehavioral scores, relative to untreated MCAO rats. Treatment with NTE increased the expression levels of SLC7A11, GPX4 and GSH, and the number of Nissl bodies in the MCAO rats.@*CONCLUSION@#Taken together, our data suggest that acute cerebral ischemia induces neuronal ferroptosis and the effects of treating MCAO rats with NTE involved inhibition of ferroptosis through the TFR1/DMT1 and SCL7A11/GPX4 pathways.
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AIM:To investigate the effects of Jiawei-Naotai formula (JWNTF) on ATF4/CHOP/Puma pathway in hippocampal neurons of ovariectomized female rats with cerebral ischemia.METHODS:The female rats were randomly divided into sham group, model group, JWNTF group and positive control group.The rats, expect in the sham group, were ovariectomized.The rats in each group were intragastric administration 11 days after ovariectomy.The rats in sham group and model group were given a gavage of 0.9％Na Cl, while the rats in other groups were administrated by corresponding therapy intragastrically for 3 d.The regional cerebral ischemia model was established by middle cerebral artery occlusion (MCAO) suture method 14 days after ovariectomy.The behaviors of the rats were evaluated 24 h after cerebral ischemia.The mRNA levels of Bax, Bcl-2 and caspase-3 were detected by RT-qPCR, and the protein expression of Bax, Bcl-2, caspase-3, ATF4, CHOP and Puma was determined by Western blot.RESULTS:Compared with sham group, the neurobehavioral scores significantly increased in other groups (P<0.05).Compared with model group, the neurobehavioral scores were significantly decreased in positive control group and JWNTF group (P<0.05).The protein expression of Bax, caspase-3, ATF4, CHOP and Puma, and the mRNA expression of Bax and caspase-3 in the hippocampus were much higher, and Bcl-2 was lower in model group than those in sham group (P<0.05).JWNTF significantly reduced the protein expression of Bax, caspase-3, ATF4 and CHOP, and the mRNA expression of Puma, Bax and caspase-3, and markedly increased the expression of Bcl-2 at mRNA and protein levels compared with model group.CONCLUSION:The JWNTF protects against brain damage induced by cerebral ischemia, which may be related to inhibitiing the expression of ATF4/CHOP/Puma pathway-related molecules at mRNA and protein levels.
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To analyze the medication features and regularity of prescriptions of Chinese medicine in treating patients with dementia based on ancient medical records. In the article, we retrieved the ancient medical records related to the treatment of dementia (from the Han Dynasty to the late Qing period) in Chinese Medical Classics, Chinese Ancient Medical Books and digital library, and then set up a medical records normalized database. The medication features and prescription rules for dementia were analyzed by frequency statistics and association rules (Apriori algorithm, improved mutual information algorithm and complex system entropy clustering). Finally, a total of 156 prescriptions were selected, involving 123 Chinese herbs, with a total frequency of 11 747 for the herbs, and 8 core prescriptions were mined. After the association rules between the frequency and prescriptions for the treatment of dementia were determined, we found that the most commonly used herbs included Fuling (Poria), Yuanzhi (Polygalae Radix), Renshen (Ginseng Radix et Rhizoma), Shichangpu (Acori Tatarinowii Rhizoma), Gancao (Glycyrrhizae Radix et Rhizoma), Danggui (Angelicae Sinensis Radix), Maidong (Ophiopogonis Radix), Baizhu (Bletillae Rhizoma), Dihuang (Rehmanniae Radix) and Ganjiang (Zingiberis Rhizoma); the frequently-used drugs compatibility was mainly for tonifying Qi-blood, regulating Yin and Yang and inducing resuscitation. The drugs were mainly of warm nature and sweet (mild) flavor, and the channel tropism of drugs mainly distributed to the heart, liver, spleen and kidney. The core prescriptions were composed of Renshen (Ginseng Radix et Rhizoma), Fuling (Poria), Yuanzhi (Polygalae Radix), Shichangpu (Acori Tatarinowii Rhizoma), and Baizhu(Bletillae Rhizoma). In conclusion, high frequency herbs and core prescriptions reflect the prescriptions by ancient physicians mainly focus on Qi-replenishing, spleen-invigorating and heart-nourishing, but also reflect the prescription rules of nourishing Yin, enriching blood, eliminating phlegm and warming Yang for the treatment of dementia. The medication features and prescription rules for the treatment of dementia obtained by association rules are useful to guide the clinical practice of Chinese medicine in treatment of dementia.
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Aim To investigate the effects of Jiawei Naotaifang on cerebral infarction area, pathological changes of brain tissue and estrogen level of focal cere-bral Iischemia in female ovariectomized rats, and cor-relation between estrogen levels and cerebral infarction area. Methods SD rats were randomly divided into sham operation group, ovariectomized group, cerebral ischemia group,model group,and drug groups(estro-gen group, Jiawei Naotaifang high dose group, Jiawei Naotaifang middle dose group, Jiawei Naotaifang low dose group). The rats in the ovariectomized group, model group, drug groups were ovariectomized, elev-enth days after the ovariectomy. The rats in the drug groups were given intragastric administration for three days. The rats in the model group, cerebral ischemia group and drug groups were prepared for cerebral is-chemia models. Neurological function scores were scored 24 hours after the success of the model, serum levels of estrogen were detected, and the brain was stained with 2, 3, 5-triphenyltetrazolium chloride (TTC) and hematoxylin-eosin staining(HE), TTC staining was used to measure the area of cerebral in-farction, and HE staining was used to observe the pathological changes of brain tissues. Results Com-pared with cerebral ischemia group,cerebral infarction area of rats in the model group increased significantly, the estrogen level was lower and the necrosis and py-knosis of cortical and hippocampus cells of rats in the model group were more obvious. Compared with model group,the cerebral infarction area of rats in the drug groups was reduced,the estrogen levels were elevated, especially in Jiawei Naotaifang high dose group and es-trogen group. The cell morphology of rats,in the estro-gen group,Jiawei Naotaifang high dose group and mid-dle dose group, was improved obviously. Cerebral in-farction area was negatively correlated with the level of estrogen. Conclusions The cerebral infarction area of cerebral ischemia in female ovariectomized rat is signif-icantly correlated with the level of estrogen. Jiawei Naotaifang can reduce the damage and alleviate brain injury of cerebral ischemia in female ovariectomized rats,which may be related to the improvement of estro-gen level.
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Objective:To provide experimental evidences for choosing murine models in the pathogenesis research of thymic impairment induced by viral infection,we compared the impacts of polycytidylic acid(Poly(I:C)) and dexamethasone(DEX) on the thymic morphology and thymic output function,and explored the implication of RLR signaling pathway.Methods: 24 male C57BL/6 mice were randomly assigned into three groups and treated with Poly(I:C),DEX,or saline respectively.Thereafter,their thymic morphology,pathological changes,thymic index,and thymic pathology were examined.Their contents of T-cell receptor excision circles (TRECs) and proportions of the naive CD4+T cell in the peripheral blood were determined to evaluate their thymic output function.The expression levels of thymic RLR/MAVS/IFN-α/β signaling pathway and IL-1β were also measured.Results: Both Poly (I:C) and DEX treatment caused thymic atrophy in appearance and structural destruction under the microscope inspection,and DEX treatment did much more severe damage,especially to the thymic cortex.TRECs decreased significantly in both groups.The proportions of na?ve/memory CD4+T cell subsets remained stable,though total CD4+T cell decreased in DEX group,while the proportion of na?ve CD4+T cell in Poly (I:C) group increased significantly.The expression of RIG-Ⅰ,MDA5,LGP2,and IFN-α/β were up-regulated in DEX group, while it remained unchanged in Poly (I:C) group.Conclusion:Both Poly (I:C) and DEX induced thymic atrophy and the impaired thymic output function.Nevertheless,the expression of RLR-IFN signaling pathway up-regulated more significantly in DEX group instead of in Poly (I:C) group.These results implied the existence of different pathological manifestations and mechanisms underlying the impaired thymic function in different animal models,as well as impact on na?ve/memory CD4+T cell proportions.Our research provides references for choosing animal models in the basic research and drug development for viral infection induced thymic atrophy based on the RLR signaling pathway.
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Objective To investigate the effect of Jiawei Naotaifang on neuronal apoptosis and the mechanism in ovariectomized rats with cerebral ischemia. Methods Female Sprague-Dawley rats(n=40)were randomly divided into sham group(n=10),model group(n=10),es-trogen group(n=10)and Jiawei Naotaifang group(n=10).The model group,estrogen group and Jiawei Naotai-fang group were ovariectomized.Eleven days after ovariectomy,the estrogen group and Jiawei Naotaifang group were given estrogen and Jiawei Naotaifang respectively intragastrically for three days.14 days after ovariecto-my,the model group,estrogen group and Jiawei Naotaifang group were modeled cerebral ischemia with Langa's method.24 hours after modeling,the apoptosis rate of neurons was detected with TUNEL,and the activation of extracellular signal-regulated kinase 1/2(ERK1/2)and c-Jun N-terminal kinase(p-JNK)in hippocampus were de-tected with Western blotting. Results Compared with the model group, the apoptosis rates decreased in Jiawei Naotaifang group and the estrogen group(P<0.001),with more activation of ERK1/2(P<0.01)and less activation of JNK(P<0.01). Conclusion Jiawei Naotaifang can protect neuron from apoptosis by promoting the activation of ERK1/2 and inhibiting the activation of p-JNK.
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<p><b>OBJECTIVE</b>To observe the therapeutic angiogenesis effect of naotai recipe (NR) on local ischemia/reperfusion (I/R) injury of rats by observing signaling pathway of hypoxia-inducible factor-lα (HIF-1α) and vascular endothelial growth factor (VEGF).</p><p><b>METHODS</b>Totally 120 Sprague-Dawley (SD) rats were randomly divided into 4 groups, namely, the normal control group (n =12), the sham-operation group (n =12), the I/R model group (n =48), and the NR group (n =48). Cerebral I/R injury models were established using thread suture method. Rats in the I/R model group and the NR group were sub-divided into 4 sub-groups according to the 1st, 3rd, 5th, and 7th I/R day (n =12). The phenomenon of neovasculization was observed by immunofluorescence staining. The protein and mRNA expression levels of HIF-la, VEGF-A, and VEGFR II receptor were detected by RT-PCR.</p><p><b>RESULTS</b>There were a large amount of labels for neovasculization in the ischemic area of the NR group. Double-immunofluorescence labeling [vWF (red) and BrdU (green)] was observed in the NR group. Compared with the model group, the HIF-1α protein expression was obviously enhanced on the 1 st day of I/R (P <0.01), and the VEGF protein expression started to enhance on the 3rd day in the NR group (P <0.01). The VEGFR protein expression level was the highest in the NR group on the 5th day of I/R (P <0.01). The protein expression of VEGF and HIF-1α started to decrease on the 7th day of I/R.</p><p><b>CONCLUSION</b>NR could strengthen angiogenesis after I/R by elevating the expression of HIF-lα and activating HIF-lα/VEGF signaling pathway.</p>
Subject(s)
Animals , Brain Ischemia , Metabolism , Cerebral Infarction , Hypoxia-Inducible Factor 1, alpha Subunit , Genetics , Metabolism , Hypoxia-Ischemia, Brain , Metabolism , Ischemia , Neovascularization, Pathologic , Rats, Sprague-Dawley , Reperfusion Injury , Signal Transduction , Vascular Endothelial Growth Factor AABSTRACT
<p><b>OBJECTIVE</b>A new rat model of cerebral infarction was developed to elucidate the contribution of vascular endothelial cell during focal cerebral infarction formation.</p><p><b>METHODS</b>Forty-eight Sprague-Dawley (SD) rats were randomly divided into the model group, sham operation group, and control group for indexes observation of triphenyltetrazolium chloride (TTC) dyeing, neurological deficit, plasma tissue-type plasminogen activator (tPA) activity, plasminogen activator inhibitor (PAI) activity, thromboxane B(2) (TXB(2)) content, and 6-keto-prostaglandin (6-keto-PGF(1alpha)) content.</p><p><b>RESULTS</b>(1) The highest neurological score appeared at 6 h after operation, descending significantly at sequential time. (2) Using TTC dyeing and optical microscope technique, pathological changes in brains were observed. (3) Compared with control group and sham operation groups, there was a decrease in tPA activity of model rats at the initial 12 h after injection of sodium laurate (P < 0.05), PAI activity decreased markedly in the model group at 24 h after injection of sodium laurate. (4) In plasma TXB(2) concentration reached the highest level compared at 6 h after injection of sodium laurate, but there were not obvious differences in plasma 6-keto-PGF(1alpha) concentration among all groups (P > 0.05).</p><p><b>CONCLUSION</b>Focal cerebral infarction in rats could be induced by some sodium laurate, showing ischemic cerebrum necrosis, function disorder of vascular endothelium-platelet, fibrinolysis abnormality. This model could play an important role in researching the contribution of vascular endothelial cell during cerebral infarction development, preventing and curing by traditional Chinese medicine.</p>