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This study aims to investigate whether tetramethylpyrazine(TMP) can stimulate angiogenesis in cerebral microvascular endothelial cells and alleviate cerebral ischemic stroke(CIS) and to explore the underlying mechanisms. In the animal study, adult Sprague-Dawley rats(n=15) were assigned into sham surgery(sham), middle cerebral artery occlusion/reperfusion(MCAO/R), and MCAO/R+TMP(intraperitoneal injection of 20 mg·kg~(-1)) groups. The neurological function was evaluated by the Z-Longa method. The cerebral infarction volume was detected by TTC staining. Enzyme-linked immunosorbent assay(ELISA) was employed to detect the expression of vascular endothelial growth factor(VEGF), angiopoietin(Ang), and platelet-derived growth factor(PDGF). Immunofluorescence staining was employed to detect Ki67 and the expression of vascular endothelial growth factor A(VEGFA) and slient information regulator 1(SIRT1). Western blot was employed to determine the expression levels of VEGFA, SIRT1, angiopoietin-2(Ang-2), and platelet-derived growth factor B(PDGFB). In the cell study, mouse brain-derived endothelial cells(Bend.3) were cultured, and the optimal concentration of TMP was determined. Then, VEGF, Ang, and PDGF were detected by ELISA after the addition of cabozantinib. Western blot was employed to measure the expression of VEGFA, Ang-2, and PDGFB. Immunofluorescence staining was used to detect CD31, CD34, and Ki67, and the proliferation, migration, and tube formation ability of Bend.3 cells were observed in vitro. Western blot and immunofluorescence staining were performed to measure the expression of SIRT1 and VEGFA after addition of the SIRT1-specific inhibitor selisistat(EX-527). The results showed that compared with the sham group, the MCAO/R group had severe neurological function damage, increased infarction volume, up-regulated expression of VEGF, VEGFA, Ang, Ang-2, PDGF, and PDGFB, and down-regulated expression of Ki67 and SIRT1(P<0.01). Compared with the MCAO/R group, the MCAO/R+TMP group presented alleviated neurological function damage, reduced infarction volume, and activated expression of VEGF, VEGFA, Ang, Ang-2, PDGF, PDGFB, Ki67, and SIRT1(P<0.01). The cell experiments showed that compared with the normal group, Bend.3 cells were activated by oxygen glucose deprivation/reoxygenation(OGD/R) treatment(P<0.05, P<0.01). Compared with the OGD/R group, the OGD/R+TMP group upregulated the expression levels of VEGF, VEGFA, Ang, Ang-2, PDGF, PDGFB, SIRT1, Ki67, CD31, and CD34, enhanced the angiogenic ability of Bend.3 cells without being inhibited by BMS or EX-527(P<0.05, P<0.01, P<0.001). The results suggest that TMP can activate the SIRT1/VEGFA signaling pathway to stimulate angiogenesis and alleviate CIS injury.
Assuntos
Isquemia Encefálica , AVC Isquêmico , Pirazinas , Acidente Vascular Cerebral , Ratos , Animais , Camundongos , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Células Endoteliais/metabolismo , Isquemia Encefálica/tratamento farmacológico , Isquemia Encefálica/genética , Isquemia Encefálica/metabolismo , Ratos Sprague-Dawley , Proteínas Proto-Oncogênicas c-sis , Sirtuína 1/genética , Sirtuína 1/metabolismo , Angiogênese , Antígeno Ki-67/metabolismo , Acidente Vascular Cerebral/tratamento farmacológico , Acidente Vascular Cerebral/genética , Transdução de Sinais , Infarto da Artéria Cerebral MédiaRESUMO
PURPOSE: Endotoxin-induced acute lung injury (ALI) is a severe disease caused by an imbalanced host response to infection. It is necessary to explore novel mechanisms for the treatment of endotoxin-induced ALI. In endotoxin-induced ALI, tetramethylpyrazine (TMP) provides protection through anti-inflammatory, anti-apoptosis, and anti-pyroptosis effects. However, the mechanism of action of TMP in endotoxin-induced ALI remains unclear. Here, we aimed to determine whether TMP can protect the lungs by inhibiting Golgi stress via the Nrf2/HO-1 pathway. METHODS AND RESULTS: Using lipopolysaccharide (LPS)-stimulated C57BL/6J mice and MLE12 alveolar epithelial cells, we observed that TMP pretreatment attenuated endotoxin-induced ALI. LPS + TMP group showed lesser lung pathological damage and a lower rate of apoptotic lung cells than LPS group. Moreover, LPS + TMP group also showed decreased levels of inflammatory factors and oxidative stress damage than LPS group (P < 0.05). Additionally, LPS + TMP group presented reduced Golgi stress by increasing the Golgi matrix protein 130 (GM130), Golgi apparatus Ca2+/Mn2+ ATPases (ATP2C1), and Golgin97 expression while decreasing the Golgi phosphoprotein 3 (GOLPH3) expression than LPS group (P < 0.05). Furthermore, TMP pretreatment promoted Nrf2 and HO-1 expression (P < 0.05). Nrf2-knockout mice or Nrf2 siRNA-transfected MLE12 cells were pretreated with TMP to explore how the Nrf2/HO-1 pathway affected TMP-mediated Golgi stress in endotoxin-induced ALI models. We observed that Nrf2 gene silencing partially reversed the alleviating effect of Golgi stress and the pulmonary protective effect of TMP. CONCLUSION: Our findings showed that TMP therapy reduced endotoxin-induced ALI by suppressing Golgi stress via the Nrf2/HO-1 signaling pathway in vivo and in vitro.
Assuntos
Lesão Pulmonar Aguda , Pirazinas , Animais , Camundongos , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/metabolismo , Antioxidantes/metabolismo , Complexo de Golgi/metabolismo , Complexo de Golgi/patologia , Heme Oxigenase-1/genética , Lipopolissacarídeos/toxicidade , Pulmão/patologia , Camundongos Endogâmicos C57BL , Fator 2 Relacionado a NF-E2/genética , Estresse Oxidativo , Transdução de Sinais , Pirazinas/farmacologiaRESUMO
Hepatic ischemia/reperfusion injury (IRI) is an adverse effect for liver transplantation which is characterized by immune response mediated inflammation. Recent studies report that neutrophil extracellular traps (NETs) are implicated in hepatic IRI. The aim of this study was to explore the mechanism of action of tetramethylpyrazine (TMP), the main chemical composition of Ligusticum chuanxiong in treatment of ischemic related diseases. Data showed that hepatic IRI increases the leak of alanine aminotransferase (ALT) and aspartate transaminase (AST), and stimulates formation of NETs. Extracellular DNA/NETs assay, hematoxylin-eosin (HE) staining, immunofluorescence assay, terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) and Western blot assay, showed that TMP significantly reduces formation of NETs and alleviates hepatic IRI. Moreover, TMP and Diphenyleneiodonium (DPI) suppressed ROS production in neutrophils. In addition, analysis showed that activation of NADPH oxidase plays a role in formation of NETs triggered by hepatic IRI. Notably, TMP inhibited formation of NETs though inhibition of NADPH oxidase. Additionally, Combination treatment using TMP and DPI was more effective compared with monotherapy of either of the two drugs. These findings show that combination therapy using TMP and DPI is a promising method for treatment hepatic IRI.
Assuntos
Antioxidantes/farmacologia , Armadilhas Extracelulares/efeitos dos fármacos , Transplante de Fígado/reabilitação , Oniocompostos/farmacologia , Pirazinas/farmacologia , Traumatismo por Reperfusão/tratamento farmacológico , Alanina Transaminase/metabolismo , Animais , Aspartato Aminotransferases/metabolismo , Ácidos Nucleicos Livres/antagonistas & inibidores , Ácidos Nucleicos Livres/sangue , Modelos Animais de Doenças , Sinergismo Farmacológico , Quimioterapia Combinada/métodos , Armadilhas Extracelulares/metabolismo , Marcação In Situ das Extremidades Cortadas , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/cirurgia , Transplante de Fígado/métodos , Masculino , NADPH Oxidases/antagonistas & inibidores , NADPH Oxidases/metabolismo , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Neutrófilos/patologia , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia , Traumatismo por Reperfusão/cirurgia , Resultado do TratamentoRESUMO
Tetramethylpyrazine (TMP, also known as Ligustrazine), which is isolated from Chinese Herb Medicine Ligustium wollichii Franchat (Chuan Xiong), has been widely used in China for the treatment of ischemic stroke by Chinese herbalists. Brain microvascular endothelial cells (BMECs) are the integral parts of the blood-brain barrier (BBB), protecting BMECs against oxygen-glucose deprivation (OGD) which is important for the treatment of ischemic stroke. Here, we investigated the protective mechanisms of TMP, focusing on OGD-injured BMECs and the Rho/Rho-kinase (Rho-associated kinases, ROCK) signaling pathway. The model of OGD-injured BMECs was established in this study. BMECs were identified by von Willebrand factor III staining and exposed to fasudil, or TMP at different concentrations (14.3, 28.6, 57.3 µM) for 2 h before 24 h of OGD injury. The effect of each treatment was examined by cell viability assays, measurement of intracellular reactive oxygen species (ROS), and transendothelial electric resistance and western blot analysis (caspase-3, endothelial nitric oxide synthase (eNOS), RhoA, Rac1). Our results show that TMP significantly attenuated apoptosis and the permeability of BMECs induced by OGD. In addition, TMP could notably down-regulate the characteristic proteins in Rho/ROCK signaling pathway such as RhoA and Rac1, which triggered abnormal changes of eNOS and ROS, respectively. Altogether, our results show that TMP has a strong protective effect against OGD-induced BMECs injury and suggest that the mechanism might be related to the inhibition of the Rho/ROCK signaling pathway.
Assuntos
Barreira Hematoencefálica/efeitos dos fármacos , Medicamentos de Ervas Chinesas/farmacologia , Células Endoteliais/metabolismo , Microvasos/efeitos dos fármacos , Pirazinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Quinases Associadas a rho/metabolismo , Animais , Barreira Hematoencefálica/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Células Cultivadas , Células Endoteliais/efeitos dos fármacos , Glucose/metabolismo , Masculino , Microvasos/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Oxigênio/metabolismo , Substâncias Protetoras/farmacologia , Ratos Sprague-DawleyRESUMO
Tetramethylpyrazine (TMP) was extracted from Ligusticum chuanxiong hort. The compound is known to have a variety of medicinal functions; in particular, it is used for the treatment of cerebral ischemic diseases. TMP-loaded hydrogels offer an excellent preparation with the capacity to bypass the blood-brain barrier, allowing treatment of the brain through intranasal administration. We prepared TMP-loaded hydrogels using carbomer 940 and evaluated the release of TMP from the hydrogel. We determined the release rate using Franz-type diffusion cell experiments with a subcutaneous-mucous-membrane model and also by a molecular dynamics (MD) simulation. In general, the former method was more complicated than the latter was. The dynamic behavior of TMP release from the hydrogel was revealed by analysis of the mean square displacement of the trajectory in the MD simulation. The coefficient of TMP diffusion from the hydrogel was calculated at different temperatures (277, 298, and 310 K) by using MD software. The results showed that the coefficient of diffusion increased with an increase in temperature. This trend was observed both experimentally and in the MD simulation. Therefore, the MD simulation was a complementary method to verify the experimental data.
Assuntos
Simulação de Dinâmica Molecular , Pirazinas/administração & dosagem , Animais , Difusão , Liberação Controlada de Fármacos , Hidrogéis/química , Ligusticum/química , Camundongos , Camundongos Endogâmicos ICR , Mucosa/metabolismo , Pirazinas/química , Pele/metabolismoRESUMO
Our recent study demonstrated that sodium arsenite at a clinically relevant dose induced nephrotoxicity in human renal proximal tubular epithelial cell line HK-2, which could be inhibited by natural product 2,3,5,6-tetramethylpyrazine (TMP) with antioxidant activity. The present study demonstrated that arsenic exposure resulted in protein and enzymatic induction of heme oxygenase-1 (HO-1) in dose- and time-dependent manners in HK-2 cells. Blocking HO-1 enzymatic activity by zinc protoporphyrin (ZnPP) augmented arsenic-induced apoptosis, ROS production and mitochondrial dysfunction, suggesting a critical role for HO-1 as a renal protectant in this procession. On the other hand, TMP, upstream of HO-1, inhibited arsenic-induced ROS production and ROS-dependent HO-1 expression. TMP also prevented mitochondria dysfunction and suppressed activation of the intrinsic apoptotic pathway in HK-2 cells. Our results revealed that the regulation of arsenic-induced HO-1 expression was performed through multiple ROS-dependent signal pathways and the corresponding transcription factors, including p38 MAPK and JNK (but not ERK), AP-1, Nrf2 and NF-κB. TMP inhibited arsenic-induced activations of JNK, p38 MAPK, ERK, AP-1 and Nrf2 and block HO-1 protein expression. The present study, furthermore, demonstrated arsenic-induced expression of arsenic response protein 2 (ARS2) that was regulated by p38 MAPK, ERK and NF-κB. To our knowledge, this is the first report showing that ARS2 involved in arsenic-induced nephrotoxicity, while TMP pretreatment prevented such an up-regulation of ARS2 in HK-2 cells. Given ARS2 and HO-1 sharing the similar regulation mechanism, we speculated that ARS2 might also mediate cell survival in this procession. In summary, our study highlighted a role of HO-1 in the protection against arsenic-induced cytotoxicity downstream from the primary targets of TMP and further indicated that TMP may be used as a potential therapeutic agent in the treatment of arsenic-induced nephrotoxicity.
Assuntos
Antioxidantes/farmacologia , Arsenitos/toxicidade , Heme Oxigenase-1/metabolismo , Túbulos Renais Proximais/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Proteínas Nucleares/metabolismo , Pirazinas/farmacologia , Compostos de Sódio/toxicidade , Fator de Transcrição AP-1/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular , Citoproteção , Relação Dose-Resposta a Droga , Regulação para Baixo , Inibidores Enzimáticos/farmacologia , Heme Oxigenase-1/antagonistas & inibidores , Humanos , Túbulos Renais Proximais/enzimologia , Túbulos Renais Proximais/patologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fatores de TempoRESUMO
AIMS: A network pharmacological analysis combined with experimental validation was used to investigate the neuroprotective mechanism of the natural product Tetramethylpyrazine(TMP). BACKGROUND: Protecting neurons is critical for acute ischemic stroke treatment. Tetramethylpyrazine is a bioactive component extracted from Chuanxiong. The neuroprotective potential of TMP has been reported, but a systematic analysis of its mechanism has not been performed. OBJECTIVE: Based on the hints of network pharmacology and bioinformatics analysis, the mechanism by which TMP alleviates oxygen-glucose deprivation-induced neuronal damage through inhibition of the HIF-1α/BNIP3 pathway was verified. METHOD: In this study, we initially used network pharmacology and bioinformatics analyses to elucidate the mechanisms involved in TMP's predictive targets on a system level. The HIF-1α/BNIP3 pathway mediating the cellular response to hypoxia and apoptosis was considered worthy of focus in the bioinformatic analysis. An oxygen-glucose deprivation (OGD)-induced PC12 cell injury model was established for functional and mechanical validation. Cell viability, lactate dehydrogenase leakage, intracellular reactive oxygen species, percentage of apoptotic cells, and Caspase-3 activity were determined to assess the TMP's protective effects. Transfection with siRNA/HIF-1α or pcDNA/HIF-1α plasmids to silence or overexpress hypoxia-inducible factor 1α(HIF-1α). The role of HIF-1α in OGD-injured cells was observed first. After that, TMP's regulation of the HIF-1α/BNIP3 pathway was investigated. The pcDNA3.1/HIF-1α-positive plasmids were applied in rescue experiments. RESULT: The results showed that TMP dose-dependently attenuated OGD-induced cell injury. The expression levels of HIF-1α, BNIP3, and the Bax/Bcl-2 increased significantly with increasing OGD duration. Overexpression of HIF-1α decreased cell viability, increased BNIP3 expression, and Bax/Bcl-2 ratio; siRNA-HIF-1α showed the opposite effect. TMP treatment suppressed HIF-1α, BNIP3 expression, and the Bax/Bcl-2 ratio and was reversed by HIF-1α overexpression. CONCLUSION: Our study shows that TMP protects OGD-damaged PC12 cells by inhibiting the HIF-1α/BNIP3 pathway, which provides new insights into the mechanism of TMP and its neuroprotective potential.
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Endoplasmic reticulum (ER) stress is a common threat to photoreceptors during the pathogenesis of chronic retinopathies and often results in irreversible visual impairment. 2,3,5,6-Tetramethylpyrazine (TMP), which possesses many beneficial pharmacological activities, is a potential drug that could be used to protect photoreceptors. In the present study, we found that the cellular growth rate of 661 W cells cultured under low glucose conditions was lower than that of control cells, while the G2/M phase of the cell cycle was longer. We further found that the mitochondrial membrane potential (ΔΨm) was lower and that ER stress factor expression was increased in 661 W cells cultured under low glucose conditions. TMP reversed these trends. Visual function and cell counts in the outer nuclear layer (ONL) were low and the TUNEL-positive rate in the ONL was high in a C3H mouse model of spontaneous retinal degeneration. Similarly, visual function was decreased, and the TUNEL-positive rate in the ONL was increased in fasted C57/BL6j mice compared with control mice. On the other hand, ER stress factor expression was found to be increased in the retinas of both mouse models, as shown by reverse transcription real-time PCR (RT-qPCR) and western blotting. TMP reversed the physiological and molecular biological variations observed in both mouse models, and ATF4 expression was enhanced again. Further investigation by using western blotting illustrated that the proportion of insoluble prion protein (PRP) versus soluble PRP was reduced both in vitro and in vivo. Taken together, these results suggest that TMP increased the functions of photoreceptors by alleviating ER stress in vitro and in vivo, and the intrinsic mechanism was the ATF4-mediated inhibition of PRP aggregation. TMP may potentially be used clinically as a therapeutic agent to attenuate the functional loss of photoreceptors during the pathogenesis of chronic retinopathies. KEY MESSAGES: ⢠Already known: TMP is a beneficial drug mainly used in clinic to enhance organ functions, and the intrinsic mechanism is still worthy of exploring. ⢠New in the study: We discovered that TMP ameliorated retinal photoreceptors function via ER stress alleviation, which was promoted by ATF4-mediated inhibition of PRP aggregation. ⢠Application prospect: In prospective clinical practices, TMP may potentially be used in the clinic as a therapeutic agent to attenuate the photoreceptors functional reduction in chronic retinopathies.
Assuntos
Fator 4 Ativador da Transcrição/fisiologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Proteínas Priônicas/efeitos dos fármacos , Agregados Proteicos/efeitos dos fármacos , Agregação Patológica de Proteínas/prevenção & controle , Pirazinas/farmacologia , Células Fotorreceptoras Retinianas Cones/efeitos dos fármacos , Degeneração Retiniana/prevenção & controle , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Eletrorretinografia , Proteínas do Olho/biossíntese , Proteínas do Olho/genética , Jejum , Feminino , Glucose/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Proteínas Priônicas/química , Agregação Patológica de Proteínas/metabolismo , Retina/metabolismo , Células Fotorreceptoras Retinianas Cones/metabolismo , Degeneração Retiniana/fisiopatologia , Método Simples-Cego , Solubilidade , Organismos Livres de Patógenos Específicos , Transcrição Gênica/efeitos dos fármacosRESUMO
The aim of this study was to investigate the therapeutic effects and underlying mechanism of tetramethylpyrazine (TMP) on lung development using a rat model of congenital diaphragmatic hernia (CDH). Nitrofen was used to induce CDH. Pregnant rats were divided into three groups: control, CDH, and CDH+TMP. In the CDH and CDH+TMP, fetuses only with left diaphragmatic hernias were chosen for analysis. Lung and body weight were recorded and lung histologic evaluations, image analysis, and western blot analysis of YAP, p-YAP and LATS1 were performed after lung processing. A markedly abnormal structure was observed, as evidenced by pulmonary hypoplasia and vascular remodeling, in the CHD. These abnormalities were improved in the CDH+TMP. There were significant differences between the CHD and CHD+TMP in percentage of medial wall thickness, arteriole muscularization, radial alveolar counts, AA%, and alveolar septal thickness. YAP expression was markedly increased in the CHD compared to the controls, which was not affected by antenatal TMP administration. However, prenatal TMP intervention significantly increased expression of LATS1 and phosphorylation of YAP in the CDH fetuses. Our results demonstrate that antenatal TMP administration improved vascular remodeling and promoted lung development in a rat model of CHD, potentially through increasing expression of LATS1 and phosphorylation of YAP.
RESUMO
BACKGROUND: Restenosis remains a clinical problem; drug-coated balloons (DCBs) have demonstrated high efficiency in this situation. DCBs prevent neointimal hyperplasia by inhibiting cell proliferation and migration. Tetramethylpyrazine (TMP) is a traditional Chinese medicine originally isolated from the rhizome of Ligusticum Walliichii, which can inhibit platelet aggregation and smooth muscle cell proliferation. We hypothesized that TMP-coated balloons (TCB) could reduce neointimal hyperplasia through the NF-κB signalling pathway. METHODS: Twenty-one New-Zealand White rabbits (2.5-3.0 kg, male) were fed high-fat diets; 36 bilateral iliac artery stenosis models were successfully established by balloon straining. Rabbits were randomly treated with TCB (n=20) or plain balloons (PBA, n=16) (3 died during model construction). Angiographies were recorded at baseline, the immediate period, and 4 weeks later. Animals were euthanized and arteries collected for histological analysis and immunohistochemical staining. Protein expression of proliferating cell nuclear antigen (PCNA) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) p65 of the vessel samples were analyzed using Western blotting. RESULTS: No difference existed in the baseline lesion characteristics or procedural results. Angiographic follow-up was successfully performed on 18 rabbits (TCB: n=20, PBA: n=16), except for 3 deaths related to the operation. Treatment with TCB was superior to that with PBA, with lower late lumen loss (0.45±0.23 vs. 0.84±0.17 mm, P<0.01). Pathological analysis confirmed the efficiency of TCB through decreasing the area stenosis rate compared with PBA (46.48%±8.22% vs. 75.24%±6.10%, P<0.01). As determined by Western blotting, significant reductions occurred in PCNA and NF-κB p65 protein intensity in the TCB group versus the PBA group (all P<0.01). TCB efficiently mitigated restenosis in the rabbit iliac artery model. CONCLUSIONS: This study elucidated that TCB could restrain intimal hyperplasia of vessels by inhibiting the activation of the NF-κB pathway to reduce inflammatory response and decrease the rate of cell proliferation through suppressing PCNA expression.
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Tetramethylpyrazine (TMP) has been studied in depth and is widely used in the treatment of many kinds of diseases in China. However, whether it has neuroprotective effects on cerebral ischemia remains unclear. An ischemia/reperfusion (I/R) injury animal model was established via middle cerebral artery occlusion in this study. We set several different groups in which the rats were performed in different ways to explore the effects of TMP on blood-brainbarrier (BBB) disruption and determine whether TMP relieved BBB disruption through blocking the JAK/STAT signaling pathway. Our results showed that TMP could reduce the neurological functional loss, decrease the brain edema and BBB permeability, as well as increase the expression of tight junction proteins via inhibiting the activation of JAK/STAT signaling pathway. Overall, we demonstrated that TMP promoted neurological recovery after I/R injury via restoring the integrity and function of BBB.
Assuntos
Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Pirazinas/farmacologia , Traumatismo por Reperfusão/patologia , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Barreira Hematoencefálica/patologia , Infarto Encefálico/complicações , Citoproteção/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/patologia , Ratos , Traumatismo por Reperfusão/complicações , Traumatismo por Reperfusão/metabolismo , Tirfostinas/farmacologia , Água/metabolismoRESUMO
In the present study, we investigated the underlying mechanism of tetramethylpyrazine (TMP)-medicated inhibition of corneal neovascularization (CNV). Our data showed that TMP could effectively downregulate the expression levels of CXCR4 mRNA and protein, as well as inhibit HUVECs, endothelial cells, tubule formation in vitro. In vivo, alkali burn (1 M NaOH) could remarkably upregulate CXCR4 expression and increase the migration of TNF-α-positive cells to corneal stroma. TMP drops could significantly downregulate CXCR4 expression in cornea, compared to the control. However, there was no difference in the downregulation of CXCR4 between TMP and FK506, an immunosuppressive drug. Moreover, the immunofluorescent staining of CD45 showed TMP and FK506 could significantly restrain the bone marrow (BM)-derived infiltration while the F4/80 staining reflects the suppression of macrophage aggregation. Meanwhile TMP could regulate the Interleukin 10 (IL-10) and FK506 could restrain the Interleukin 2 (IL-2). Furthermore, TMP and FK506 significantly ameliorate corneal opacity and neovascularization. Clinical assessment detected an obvious improvement in TMP and FK506 treatment groups, compared to controls in vivo. Thus, TMP had similar effects in inhibition of immune response and CNV by suppressing BM-infiltrating cells into cornea as FK506. TMP could be a potential agent in eye-drop therapy for cornea damaged by Alkali Burn.
Assuntos
Álcalis/efeitos adversos , Queimaduras Químicas/tratamento farmacológico , Córnea/efeitos dos fármacos , Neovascularização da Córnea/tratamento farmacológico , Neovascularização Patológica/tratamento farmacológico , Pirazinas/farmacologia , Animais , Queimaduras Químicas/metabolismo , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Córnea/metabolismo , Neovascularização da Córnea/metabolismo , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Neovascularização Patológica/metabolismo , Receptores CXCR4/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacosRESUMO
Tetramethylpyrazine (TMP) is a natural product extracted from Rhizoma Chuanxiong. Liposomes are an excellent delivery method that is suitable for the penetration of the blood-brain barrier (BBB). TMP-loaded liposomes are suitable for the treatment of cerebral ischemic diseases. We measured the morphology, particle size, zeta-potential, entrapment efficiency, and content of TMP-loaded liposome. The electrochemical method was adopted to determine entrapment efficiency for the first time. In addition, their diffusion through BBB models, which were built not only by in vitro cell culture but also by the ex vivo mouse subcutaneous-mucous-membrane (mSMM) method, was explored. The results showed that TMP and TMP-loaded liposomes could effectively permeate both BBB models. Therefore, the study indicated that liposomes were a novel carrier that could deliver TMP across the body barrier models and then release TMP slowly.
Assuntos
Barreira Hematoencefálica/efeitos dos fármacos , Modelos Biológicos , Pirazinas/farmacologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Eletroquímica , Feminino , Lipossomos , Camundongos Endogâmicos ICR , Mucosa/efeitos dos fármacos , Tamanho da Partícula , Ratos Sprague-Dawley , Eletricidade EstáticaRESUMO
Tetramethylpyrazine (TMP) was originally isolated from a traditional Chinese herbal medicine, Ligusticum chuanxiong In the present study, TMP exhibits potent antitumor activities in vitro. However, the molecular mechanisms remain to be defined. Hence, this study aims to investigate the antiproliferative and apoptotic effects of TMP on HepG2 and elucidate the underlying mechanisms. Analyses using Cell Counting Kit-8 and real-time cell analyzer indicated that TMP significantly inhibited HepG2 cell proliferation. We also observed that TMP induced cell cycle arrest at the G0/G1 checkpoint and apoptosis, using flow cytometry and high-content screening. Furthermore, our results predicted that TMP could directly decrease mitochondrial membrane potential (Δψm), increase the release of cytochrome c, and increase caspase activation, indicating that mitochondrial pathway apoptosis could be the mechanism for TMP within HepG2 cells. Moreover, TMP altered expression of p53 and the Bcl-2/Bax protein ratio, which revealed that TMP induced cell cycle arrest and caspase-dependent mitochondrial apoptosis in HepG2 cells in vitro. These studies provided mechanistic insights into the antitumor properties of TMP, which may be explored as a potential option for treatment of hepatocellular carcinoma.
Assuntos
Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/tratamento farmacológico , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Neoplasias Hepáticas/tratamento farmacológico , Mitocôndrias/efeitos dos fármacos , Pirazinas/farmacologia , Proteína Supressora de Tumor p53/metabolismo , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Citocromos c/metabolismo , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Células Hep G2 , Humanos , Neoplasias Hepáticas/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Fase de Repouso do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Proteína X Associada a bcl-2/metabolismoRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Ligusticum species (Umbelliferae) have been widely used in traditional Chinese medicine, Korean folk medicine and Native American medicine for their medicinal and nutritional value. Decoctions of the rhizomes are used in treatment and prophylaxis of migraine, anemia and cardiovascular conditions including stroke. AIM OF STUDY: This review is intended to fully compile the constituents of locally and traditionally used Ligusticum species, present their bioactivities and highlight potential leads for future drug design, and thus, provide a reference for further research and application of these species. Emphasis is also placed on current trends in the pharmacokinetic studies of the major constituents. METHODS: The literature discussed is derived from readily accessible papers spanning the early 1990s to the end of 2015. Information was collected from journals, books and online searches (Google Scholar, PubMed, ScienceDirect, SciFinder, Springerlink and CNKI). RESULTS: The major phytoconstituents, 154 of which are presented in this review, include alkaloids, phthalides and phenolic acids. The crude extracts and isolated constituents have exhibited a wide range of in vitro and in vivo pharmacologic effects, including cardioprotective, antioxidant, anti-inflammatory and neuroprotective activities. The bioactive alkaloid tetramethylpyrazine (TMP) has attracted the most attention for its potent effect on calcium channels, anti-platelet as well as anti-inflammatory effects. Pharmacokinetic studies of major constituents have also been summarized. CONCLUSION: The pthalides, organic acids and alkaloids of Ligusticum species have emerged as a good source of traditional medicines for the management of cardio- and cerebrovascular conditions, inflammation and neurogenerative disorders. The species discussed in this review have demonstrated wide pharmacological actions and have great potential to yield multipotent drugs if challenges such as poor bioavailability, solubility and toxicological profiles are addressed. Apart from the rhizomes, pharmacological activities of other botanical parts also need to be studied further. Expansion of research to cover other species in the Ligusticum genus would provide more opportunities for the discovery of new bioactive principles.
Assuntos
Ligusticum/química , Medicina Tradicional , Compostos Fitoquímicos/uso terapêutico , FarmacocinéticaRESUMO
We focused on the production and evaluation of a modified "double-hit" induced acute lung injury (ALI) model, which closely mimics the clinical situation. Further, tetramethylpyrazine (TMP), an alkaloid contained in ligustrazine was evaluated for its potent anti-inï¬ammatory effects in this model. Rats were randomized into 4 groups: G1 (NS control group), G2 ("double-hit" group), G3 (low dosage TMP group) and G4 (high dosage TMP group). The rats in G2, G3 and G4 were intraperitoneally injected with a low dose of LPS followed by intratracheal injection with median dose of LPS to establish the "double-hit" model. The rats in G3, G4 were intraperitoneally injected with low (G3), high (G4) dosage TMP for the protection against ALI. Upon termination of the experiment, TMP attenuated the harmful changes in animal model reaction, breathing frequency, histological examination, lung W/D-weight ratio, BAL fluid PMNs percentage, MPO activity and ROCK2 mRNA expression. We found inhibiting RhoA/ROCK pathway might attribute to TMP-induced protection against ALI.