【Abstract】ObjectiveTo explore the protective effect of ligustrazine on the ischemia-reperfusion injury of rat liver. MethodsNinety-six healthy SD rats were divided randomly into three groups: sham operation group, ischemiareperfusion group（I/R group) and ischemia plus ligustrazine reperfusion group(therapy group).The plasm ALT,AST and LDH were measured before operation,at thirty minutes,six hours and twentyfour hours after operation. One week survival and liver pathological change of every group were observed, and the hepatocyte apoptosis index was measured simultaneously.ResultsOne week survival of therapy group was higher than that of I/R group (P＜0.05). The plasm ALT,AST and LDH of therapy group and I/R group were higher than those of the sham operation group significantly (P＜0.05), and those of therapy group were lower than those of the I/R group (P＜0.05). Light microscopy indicated that the liver sinusoid and central veins were congested remarkably after operation, the hepatocyte necrosis in I/R group was more severe than that in therapy group, and the hepatocyte apoptosis index of I/R group was higher than that of therapy group (P＜0.05). ConclusionThe protective effect of ligustrazine on ischemia-reperfusion injury of rat liver is obvious.
Objective To evaluate the efficacy and safety of tetramethylpyrazine (TMP) for diabetic peripheral neuropathy (DPN). Methods Randomized controlled trials were identified from CBM (1978-2003.3), TCMLRS (1980-2003.3), Medline (1970-2003.3), EMbase (1970-2003.3) and Cochrane Library (issue 3, 2003). We handsearched Journal of Traditional Chinese Medicine (1990-2002), New Chinese Medicine (1990-2002), Chinese Journal of Integrated Traditional and Western Medicine (1990-2002) and Research of Traditional Chinese Medicine (1990-2002). Papers of the controlled trials of clinical therapeutic studies on DPN treatment by Chinese medicine herb TMP were included and analyzed according to the criteria of the Cochrane Handbook in 1997. Results Six RCTs involving 669 patients were included, with all trials of low methodological quality. Meta-analysis indicated TMP was more effective than western medicine on pain or numbness of extremities of DPN [The pooled OR = 10.12, 95%CI (6.70 to15.28), P=0.000] and motor nerve conduction velocity change of common peroneal nerves and median nerves . Only one trial reported the side effects of TMP, such as dizziness and headache. Conclusions Based on the review, TMP infusion may have positive effect on DPN. However, the evidence is not b enough due to the general low methodological quality, so we can’t draw a reliable conclusion about the effects of TMP for DPN at the moment. Further large randomized, double blind, placebo-controlled trial are needed.
In order to study the mechanism of the inhibitory effect of salvia miltiorrhiza (SM) and tetramethyl pyrazine (TP) on scar fibroblast, the DNA content of fibroblast and the all distribution in cellular cycle was measured by FCM. The hypertrophic scar tissue of chest was chosen for primary culture of fibroblast. Then this cultured cell was reacted with SM and TP. FCM was used to measure the DNA index and duration of cellular cycle. The results showed that: 1. SM and TP had little effect on DNA index, but when the concentration of drugs reached the threshold, they could increase the amount of fibroblasts in C2-M stage and the duration of G2-M stage was prolonged; 2. TP could also prolong the duration of S-stage; 3. SM and TP could prolong the multiplication time of fibroblasts and this effect was correlated postively with the dosage of drug. The conclusions were that the inhibitory effect of SM was the result of inhibiting the mitosis of cells and the cellular cycle be at a standstill in G2-M stage. The inhibitory effect of TP was due to the inhibition of synthesis and duplication of DNA and cellular mitosis, and the cellular cycle was also at a standstill in G2-M stage.
Objective To provide evidence for clinical practice by assessing the effectiveness and safety of Ligustrazine for primary nephrotic syndrome. Methods We searched MEDLINE (1966.1-2002.12), EMBASE (1975-2002.12), CBM (1979.1-2002.12), Chinese Evidence-Based Medicine/Cochrane Centre Database (CEBM/CCD, Issue 4, 2002) , Cochrane Library, and SCI (1985-2002.12) and handsearched 15 kinds of journals (including Journal of Nephrology et al) (1980-2003.2) for the randomized controlled trials (RCTs).Jadad score was used to assess the quality of RCTs. The outcomes of short term and long term effectiveness, and adverse effect of the treatment were analyzed by RevMan 4.1. Results Thirteen RCTs involving 675 patients met inclusion criteria. Jadad scores of each trial was 1 point. Meta-analysis of 4 studies showed that Ligustrazine had significant better short term effect [OR 4.24, 95% confidence interval (CI) 1.76 to 10.19], lowered 24 h urine protein (OR -0.36, 95% CI -0.71 to -0.02), improved renal function [ creatinine level in children group: (OR -3.34, 95% CI -5.25 to -1.43), creatinine level in adult group: (OR -48.29, 95% CI -68.24 to -28.35)], and increased serum albumin level (OR 3.61, 95% CI 2.61 to 4.61). Whether Ligustrazine had long term side effect was not confirmed. No adequate evidence showed that Ligustrazine could reduce the relapse rate of primary nephrotic syndrome. Conclusions Meta-analysis of low quality RCTs suggest that Ligustrazine does work in primary nephritic syndrome in short term observation. No adequate evidence shows that Ligustrazine has severe side effect or can reduce the relapse rate of primary nephrotic syndrome. We can’t draw a conclusion that Ligustrazine is safe in primary nephrotic syndrome treatment.A rigorously designed, randomized, double blind, placebo controlled trial are required.
Objective To explore the mechanisms for repairing spinal cord injury (SCI) with tetramethylpyrazine-loaded electroconductive hydrogel (hereinafter referred to as “TGTP”). Mehtods A total of 72 female Sprague-Dawley rats were randomly divided into 4 groups: sham operation group (group A), SCI group (group B), SCI+electroconductive hydrogel group (group C), and SCI+TGTP group (group D). Only the vertebral plate was removed in group A, while the remaining groups were subjected to a whole transection model of spinal cord with a 2 mm gap in the lesions. The recovery of hindlimb motor function was evaluated by Basso, Beattie, Bresnahan (BBB) score and modified Rivlin-Tator inclined plate test before operation and at 1, 3, 7, 14, and 28 days after operation, respectively. Animals were sacrificed at 7 days and 28 days after modeling. Neovascularisation was observed by immunofluorescence staining of CD31 and the expression levels of angiopoietin 1 (Ang-1) and Tie-2 were assessed by Western blot assay. At 28 days postoperatively, the expression levels of pro-angiogenic related proteins, including platelet-derived growth factor B (PDGF-B), PDGF receptor β (PDGFR-β), vascular endothelial growth factor A (VEGF-A), and VEGF receptor 2 (VEGFR-2), were also assessed by Western blot. The fibrous scar in the injured area was assessed using Masson staining, while neuronal survival was observed through Nissl staining. Furthermore, LFB staining was utilized to detect myelin distribution and regeneration. Immunofluorescence and Western blot assay were employed to evaluate the expression of neurofilament 200 (NF200). Results The hindlimb motor function of rats in each group gradually recovered from the 3rd day after operation. The BBB score and climbing angle in group D were significantly higher than those in group B from 3 to 28 days after operation, and significantly higher than those in group C at 14 days and 28 days after operation (P<0.05). Masson staining showed that the collagen volume fraction in groups B-D were significantly higher than that in group A, and that in group D was significantly lower than that in groups B and C (P<0.05); a small amount of black conductive particles were scattered at the broken end in group D, and the surrounding collagen fibers were less than those in group C. Nissl and LFB staining showed that the structure of neurons and myelin sheath in the injured area of spinal cord in group D was relatively complete and continuous, and the number of Nissl bodies and the positive area of myelin sheath in group D were significantly better than those in groups B and C (P<0.05). NF200 immunofluorescence staining and Western blot assay results showed that the relative expression of NF200 protein in group D was significantly higher than that in groups B and C (P<0.05). CD31 immunofluorescence staining showed that the fluorescence intensity of group D was better than that of groups B and C at 28 days after operation, and tubular or linear neovascularization could be seen. The relative expressions of Ang-1 and Tie-2 proteins in group D were significantly higher than those in groups B and C at 7 and 28 days after operation (P<0.05). The relative expressions of PDGF-B and PDGFR-β proteins in group D were significantly higher than those in groups B and C, and group B was significantly higher than group C at 28 days after operation (P<0.05). The relative expressions of VEGF-A and VEGFR2 proteins in group D were higher than those in groups B and C, showing significant difference when compared with group B (P<0.05), but only the expression of VEGF-A protein was significantly higher than that in group C (P<0.05). There was significant difference only in VEGFR-2 protein between groups B and C (P<0.05). Conclusion TGTP may enhance the revascularization of the injured area and protect the neurons, thus alleviating the injury of spinal cord tissue structure and promoting the recovery of neurological function after SCI in rats.
目的 探讨川芎嗪对大鼠重症急性胰腺炎（SAP）脑损伤的保护作用。方法 将72只健康Wistar大鼠按数字表法随机均分为对照组、SAP组和川芎嗪治疗组3组。对照组仅剖腹翻动胰腺后即缝合腹壁； SAP组采用胰腺被膜下均匀注射5%牛磺胆酸钠（2ml/kg体重）制备SAP动物模型；川芎嗪治疗组在SAP建模后5min于大鼠尾静脉注射川芎嗪注射液（100mg/kg体重）。各组大鼠分别于术后第6、12及24 h观察胰腺组织及脑组织的病理改变，检测血清淀粉酶、 脑组织含水量和微血管内白细胞聚集附壁计数，以及脑组织中MDA、TNF-α和IL-1β水平。结果 对照组胰腺组织无明显改变；SAP组胰腺组织腺泡细胞坏死，结构不清，间质水肿，红细胞漏出，部分腺导管扩张，有点片状出血坏死，炎性细胞浸润；川芎嗪治疗组胰腺组织病理改变较同一时相的SAP组明显减轻。对照组脑组织无明显改变；SAP组脑组织神经元细胞水肿，微血管内白细胞聚集及附壁，脑组织内有炎性细胞增生、聚集，且随时间延长上述表现逐渐加重；川芎嗪治疗组脑组织病理改变较同一时相的SAP组明显减轻。SAP组大鼠各时相脑组织含水量和微血管内白细胞聚集附壁计数，脑组织中TNF-α、IL-1β和MDA水平，以及血淀粉酶含量均明显高于对照组（P＜0.05）；川芎嗪治疗组大鼠各时相的上述指标均明显低于SAP组（P＜0.05）。结论 大鼠脑组织中的TNF-α、IL-1β及MDA参与了SAP脑损伤的病理过程，川芎嗪对SAP大鼠脑损伤具有保护、治疗作用。
Objective To observe the effect of tetramethypyrazine (TMP) on the expression of hypoxia-related factors in human umbilical vein endothelial cells (HUVECs). Methods The second to fifth passage cultured HUVECs were divided into five groups: control group, CoCl2induced hypoxic group and 50, 100, 200 mu;mol/L TMP treatment groups. HUVECs in control group were not treated. HUVECs inCoCl2induced hypoxic group were treated with 150 mu;mol/LCoCl2for four hours. HUVECs in 50, 100, 200 mu;mol/L TMP treated groups were pretreated with 150 mu;mol/LCoCl2 for four hours, followed by treatment with 50, 100, 200 mu;mol/L TMP for eight hours. Real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the mRNA levels of prolyl hydroxylase 2 (PHD2), hypoxia-induced factor-1alpha;(HIF-1alpha;) and vascular endothelial growth factor (VEGF). Protein levels of PHD2, HIF-1alpha;, and VEGF were detected using Western blot. Results Compared with the control group, theCoCl2 induced hypoxic group showed decreased mRNA and protein levels of PHD2 (t=3.734, 3.122;P＜0.05), while those of HIF-1alpha; and VEGF increased (HIF-1alpha; mRNA:t=4.589,P＜0.05; HIF-1alpha; protein:t=3.778,P＜0.05. VEGF mRNA:t=3.926,P＜0.05; VEGF protein:t=3.257,P＜0.05). Compared with theCoCl2 induced hypoxic group, 50, 100, 200 mu;mol/L TMP treated groups showed increased mRNA and protein levels of PHD2 (PHD2 mRNA: t=3.286, 3.617, 3.886;P＜0.05. PHD2 protein: t=2.813, 3.026, 3.078; P＜0.05); while those of VEGF decreased (VEGF mRNA: 50 mu;mol/L TMP: t=1.696,P＞0.05; 100 mu;mol/L TMP:t=2.974,P＜0.05; 200 mu;mol/L TMP: t=3.492,P＜0.05; VEGF protein: 50 mu;mol/L TMP: t=1.986,P＞0.05; 100 mu;mol/L TMP: t=2.976,P＜0.05; 200 mu;mol/L TMP:t=3.136,P＜0.05); although changes in HIF-1alpha;mRNA levels were not statistically significant (t=1.025, 0.726, -1.386;P＞0.05), showed a decrease in HIF-1alpha;protein levels (50 mu;mol/L TMP: t=2.056,P＞0.05; 100 mu;mol/L TMP:t=3.058,P＜0.05; 200 mu;mol/L TMP:t=3.828,P＜0.05). ConclusionIn HUVECs, TMP can upregulate the mRNA and protein expression of PHD2, while down regulating HIF-1alpha; protein expression and VEGF mRNA and protein expression under acute hypoxic conditions.
Objective:To observe the intervention effect of the tetra methylpyraz ine on the rds mice with retinitis pigmentosa. Methods:A total of 84 rds mice were randomly divided into 2 groups, with 42 mice in each group. The mice in the experimental group underwent intraperitoneal cavity injection with hydrochlor i c tetramethylpyrazine (80 mg/kg, twice per day) at the date of birth and till 35 days after birth, whereas the normal saline was injected into the intraperito n eal cavity of rats in the control group. The mice were sacrificed 0, 3, 7, 14, 2 1, 28, 35 days after birth, and the eyeballs were enucleated for the routine pat hologic examination with the light microscope. The apoptosis of photoreceptor ce ll nuclei was detected by terminal deoxynucleotidyl transferasemediated dUTP n i ck endlabeling (TUNEL) technigue and the expression of bcl2 in retina was de tect by immunohistochemistry method. Results:The results of li ght microscopy s howed that the layer number of retinal photoreceptor cell nuclei with tetramethy lpyrazine treatment was increased 14, 21, 28, 35 days after the treatment compar ed with that in the control group（P＜0.01）. The results of electron-micro scope suggested that tetramethylpyrazine might reduce lesions in the photoreceptor cells and the destruction of the disc member, mitochondrion，and outer limiting me mbrane in the photoreceptor outer segment in rds mice. The apoptosis of the phot oreceptor cell nuclei reduced in rds mice 3, 7, 14, 21, 28 and 35 days after the treatment compared with that in the control group （P＜0.01）. The express ion of bcl-2 in the matrix of retinal photoreceptor cell nuclei and its inner and o u ter segments increased significantly in rds mice 3，7, 14, 21, 28 and 35 days af ter the treatment （P＜0.05）. Conclusions:Tetramethylpyra zine might reduce ret inal photoreceptor apoptosis by upregulating the expression of bcl-2 in the m at rix of retinal photoreceptor cell nuclei or its inner and outer segments in rds mice.
The aim of this research is to evaluate the effect of tetramethylpyrazine (TMP) and connective tissue growth factor (CTGF) miRNA plasmids on the expressive levels of CTGF, transforming growth factor-beta (TGF-beta) and type Ⅰ collagen of rat hepatic stellate cells (HSC) which are stimulated by high glucose. The rat HSCs which were successfully transfected rat CTGF miRNA plasmids and the rat HSCs which were successfully transfected negative plasmids were cultured in vitro. After stimulus of the TMP and the high glucose, the protein levels and gene expressive levels of CTGF, TGF-beta and type Ⅰ collagen were tested. The results indicated that high glucose increased the expression of CTGF mRNA, CTGF protein, TGF-beta mRNA,TGF-beta protein and type Ⅰ collagen (P<0.05). The expressive levels of CTGF mRNA, CTGF protein, TGF-beta mRNA, TGF-beta and type Ⅰ collagen in TMP group were lower than those in high glucose group and showed statistically significant differences (P0.05). Compared with high glucose group, the expressive levels of CTGF mRNA, CTGF protein, TGF-beta mRNA, TGF-beta and type Ⅰ collagen in rat CTGF miRNA plasmid interference group were significantly lower (P<0.05). However, no statistically significant difference was found in CTGF mRNA and CTGF protein levels between TMP group and CTGF miRNA group (P>0.05), while type Ⅰ collagen levels showed statistically significant differences (P<0.05). It is concluded that high glucose could promote the expressions of CTGF, TGF-beta and type Ⅰ collagen, and TMP and rat CTGF miRNA plasmids could reduce the expressions of CTGF, TGF-beta, type Ⅰ collagen.
Objective To investigate the neuroprotective effect of conducting hydrogel loaded with tetramethylpyrazine sustained-release microparticles (hereinafter referred to as “TGTP hydrogel”) on spinal cord injury rats. Methods Forty-eight adult female Sprague Dawley rats were randomly divided into 4 groups: sham operation group (group A), model group (group B), conductive hydrogel group (group C), and TGTP hydrogel group (group D), with 12 rats in each group. Only laminectomy was performed in group A, and complete spinal cord transection was performed in groups B, C, and D. Basso-Bettie-Bresnahan (BBB) score was used to evaluate the recovery of hind limb motor function of each group before modeling and at 1, 3, 7, 14, and 28 days after modeling, respectively. At 28 days after modeling, the rats were sacrificed for luxol fast blue (LFB) staining to detect myelin regeneration. Nissl staining was used to detect the survival of neurons. Immunohistochemical staining was used to evaluate the expression of inflammation-related factors [nuclear factor кB (NF-кB), tumor necrosis factor α (TNF-α), and interleukin 10 (IL-10)]. Immunofluorescence staining and Western blot were used to evaluate the expression of neurofilament 200 (NF200). RseultsBBB scores of group A were significantly better than those of the other three groups at all time points after modeling (P<0.05); at 14 and 28 days after modeling, there was no significant difference in BBB scores between groups C and D (P>0.05), but the BBB score of group D was significantly better than that of group B (P<0.05). LFB staining and Nissl staining showed that the structure of neurons and myelin in group A was intact, and the myelin integrity and survival number of neurons in group D were significantly better than those in groups B and C. Immunohistochemical staining showed that the absorbency (A) value of NF-кB and TNF-α in group A were significantly lower than those in groups B and C (P<0.05), the A value of IL-10 was significantly higher than that in the other three groups (P<0.05); the A value of NF-κB in group D was significantly lower than that in groups B and C, the A value of TNF-α in group D was significantly lower than that in group B, while the A value of IL-10 in group D was significantly higher than that in group B (P<0.05). Immunofluorescence staining showed that the structure of neurons and nerve fibers in group A was clear and the fluorescence intensity was high. The fluorescence intensity of NF200 in group D was higher than that in groups B and C, and some nerve fibers could be seen. Western blot analysis showed that the relative expression of NF200 in group A was the highest, and the relative expression of NF200 in group D was significantly higher than that in groups B and C (P<0.05). Conclusion The TGTP hydrogel can effectively promote the recovery of motor function in rats with spinal cord injury, and its mechanism may be related to the regulation of inflammatory response.