Impacts of national volume-based drug procurement policy on the utilization and costs of antihypertensive drugs in a Chinese medicine hospital: an interrupted time series analysis of 5138 patients.

Objectives: The study aimed to estimate the effects of National Volume-based Drug Procurement (NVBP) policy on drug utilization and medical expenditures of hypertension patients in public medical institutions in mainland China. Methods: This study used patient-level data based on electronic health records retrieved from the hospital information system of Nanjing Hospital of Chinese Medicine. Data on patients with hypertension who received care at this institution between 2016 and 2021 was used for analysis. Segmented linear regression models incorporating Interrupted Time Series (ITS) analysis were adopted to examine the effects of NVBP policy on drug utilization and health expenditures of eligible patients. Drug utilization volume and health expenditures were the primary outcomes used to assess the policy effects, and were measured using the prescription proportion of each drug class and the overall per-encounter treatment costs. Results: After the implementation of NVBP policy, the volume of non-winning drugs decreased from 54.42% to 36.25% for outpatient care and from 35.62% to 15.65% for inpatient care. The ITS analysis showed that the volume of bid-winning drugs in outpatient and inpatient settings increased by 9.55% (p < 0.001) and 6.31% (p < 0.001), respectively. The volume changes in non-volume based purchased (non-VBP) drugs differed between outpatients and inpatients. The proportion of non-VBP drugs immediately increased by 5.34% (p = 0.002) overall, and showed an upward trend in the outpatient setting specially (p < 0.001) during the post-intervention period. However, no significant differences were observed in the proportion of non-VBP drugs in inpatient setting (p > 0.05) in term of level change (p > 0.05) or trend change (p > 0.05). The average per-visit expenditures of outpatients across all drug groups exhibited an upward trend (p < 0.05) post policy intervention. In addition, a similar increase in the overall costs for chemical drugs were observed in inpatient settings (coefficient = 2,599.54, p = 0.036), with no statistically significant differences in the regression slope and level (p = 0.814). Conclusion: The usage proportion of bid-winning drugs increased significantly post policy intervention, indicating greater use of bid-winning drugs and the corresponding substitution of non-winning hypertensive drugs. Drug expenditures for outpatients and health expenditures per visit for inpatients also exhibited an upward trend, suggesting the importance of enhanced drug use management in Traditional Chinese Medicine hospital settings.

Effect and mechanism of Banxia Xiexin decoction in colorectal cancer: A network pharmacology approach.

BACKGROUND: Banxia Xiexin decoction (BXD) is a traditional Chinese medicine with anti-colorectal cancer (CRC) activity. However, its bioactive constituents and its mechanism of action remain unclear. Herein, we explored the mechanism of action of BXD against CRC using a network pharmacology approach. METHODS: First, the targets of the main chemical components of BXD were predicted and collected through a database, and the intersection of compound targets and disease targets was obtained. Subsequently, protein-protein interaction network analysis, Gene Ontology enrichment, and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis were performed to explore the potential mechanisms underlying the effects of BXD on CRC. Finally, a CRC cell model and a CRC xenograft model in nude mice were utilized to further determine the mechanism of action. RESULTS: A compound-therapeutic target network of BXD was constructed, revealing 146 cellular targets of BXD. The phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling axis was identified as the main target of BXD. Using in vitro and in vivo models, the activity of BXD against CRC was found to be mediated through ferritinophagy by targeting the PI3K/AKT/mTOR axis, leading to intracellular iron accumulation, reactive oxygen species activation, and finally ferroptosis. CONCLUSIONS: Through the application of network pharmacology and in vitro/in vivo validation experiments, we discovered that BXD exerts anti-CRC effects via the ferritinophagy pathway. Furthermore, we elucidated the potential mechanism underlying its induction of ferritinophagy. These findings demonstrate the significant potential of traditional drugs in managing CRC and support their wider clinical application in combination chemotherapy, targeted therapy, and immunotherapy.

The Efficacy and Mechanism of Qinghua Jianpi Recipe in Inhibiting Canceration of Colorectal Adenoma Based on Inflammatory Cancer Transformation.

Objective: This study is aimed at exploring the effect of Qinghua Jianpi Recipe on preventing colon polyp recurrence and inhibiting the progress of "inflammatory cancer transformation." And another goal is to explore the changes of intestinal flora structure and intestinal inflammatory (immune) microenvironment of mice with colon polyps treated by Qinghua Jianpi Recipe and to clarify its mechanism. Methods: Clinical trials were conducted to confirm the therapeutic effect of Qinghua Jianpi Recipe on patients with inflammatory bowel disease. The inhibitory effect of Qinghua Jianpi Recipe on "inflammatory cancer transformation" of colon cancer was confirmed by an adenoma canceration mouse model. Histopathological examination was used to evaluate the effects of Qinghua Jianpi Recipe on intestinal inflammatory state, adenoma number, and pathological changes of adenoma model mice. The changes of inflammatory indexes in intestinal tissue were tested by ELISA. Intestinal flora was detected by 16S rRNA high-throughput sequencing. Short-chain fatty acid metabolism in the intestine was analyzed by targeted metabolomics. Network pharmacology analysis of possible mechanism of Qinghua Jianpi Recipe on colorectal cancer was performed. Western blot was used to detect the protein expression of the related signaling pathways. Results: Qinghua Jianpi Recipe can significantly improve intestinal inflammation status and function in patients with inflammatory bowel disease. Qinghua Jianpi Recipe could significantly improve the intestinal inflammatory activity and pathological damage of adenoma model mice and reduce the number of adenoma. Qinghua Jianpi Recipe significantly increased the levels of Peptostreptococcales_Tissierellales, NK4A214_group, Romboutsia, and other intestinal flora after intervention. Meanwhile, the treatment group of Qinghua Jianpi Recipe could reverse the changes of short-chain fatty acids. Network pharmacology analysis and experimental studies showed that Qinghua Jianpi Recipe inhibited the "inflammatory cancer transformation" of colon cancer by regulating intestinal barrier function-related proteins, inflammatory and immune-related signaling pathways, and free fatty acid receptor 2 (FFAR2). Conclusion: Qinghua Jianpi Recipe can improve the intestinal inflammatory activity and pathological damage of patient and adenoma cancer model mice. And its mechanism is related to the regulation of intestinal flora structure and abundance, short-chain fatty acid metabolism, intestinal barrier function, and inflammatory pathways.

Qingchang Wenzhong Decoction reduce ulcerative colitis in mice by inhibiting Th17 lymphocyte differentiation.

BACKGROUND: Qingchang Wenzhong Decoction (QCWZD), a chinese herbal prescription, is widely used for ulcerative colitis (UC). Nevertheless, the active ingredients and mechanism of QCWZD in UC have not yet been explained clearly. PURPOSE: This research focuses on the identification of the effective ingredients of QCWZD and the prediction and verification of their potential targets. METHODS: The UC mice were established by adding 3.0% dextran sulfate sodium (DSS) to sterile water for one week. Concurrently, mice in the treatment group were gavage QCWZD or mesalazine. LC-MS analyzed the main components absorbed after QCWZD treatment, and network pharmacology predicted their possible targets. ELISA, qPCR, immunohistochemistry and immunofluorescence experiments were used to evaluate the colonic inflammation level and the intestinal barrier completeness. The percentage of Th17 and Treg lymphocytes was detected by flow cytometry. RESULTS: After QCWZD treatment, twenty-seven compounds were identified from the serum. In addition, QCWZD treatment significantly reduced the increased myeloperoxidase (MPO) and inflammatory cell infiltration caused by DSS in the colonic. In addition, QCWZD can reduce the secretion of inflammatory factors in serum and promote the expression of mRNAs and proteins of occludin and ZO-1. Network pharmacology analysis indicated that inhibiting IL-6-STAT3 pathway may be necessary for QCWZD to treat UC. Flow cytometry analysis showed that QCWZD can restore the normal proportion of Th17 lymphocytes in UC mice. Mechanistically, QCWZD inhibited the phosphorylation of JAK2-STAT3 pathway, reducing the transcriptional activation of RORγT and IL-17A. CONCLUSIONS: Overall, for the first time, our work revealed the components of QCWZD absorbed into blood, indicated that the effective ingredients of QCWZD may inhibit IL-6-STAT3 pathway and inhibit the differentiation of Th17 lymphocytes to reduce colon inflammation.

Effect of Glycyrrhiza uralensis against ulcerative colitis through regulating the signaling pathway of FXR/P-gp.

OBJECTIVE: Ulcerative colitis (UC) is a moderate to severe inflammatory bowel disease, with a characteristic inflammatory response. Chinese herbal medicine can play a role in UC treatment. Herein, we aimed to investigate the function of Glycyrrhiza uralensis in UC treatment and the underlying mechanism. METHODS: After establishing an animal model of UC, different agents of kuijieguanchang prescription, Glycyrrhiza uralensis, mesalazine, and GW4064 were administrated to mice. The apoptosis rate was measured by TUNEL assay, and the expression of different biomarkers was tested by western blot and qPCR. RESULTS: Glycyrrhiza uralensis could regulate apoptosis of intestinal mucosal cells, through regulating the expression of apoptosis-related proteins and protective proteins of intestinal mucosa. The administration of Glycyrrhiza uralensis could greatly enhance the expression of muc1, muc3, and the pro-apoptotic protein, BAX. The proteins involved in malignancy from UC, such as Bcl-2 and fgf-15, were dramatically downregulated after using the Glycyrrhiza uralensis. Moreover, it was illustrated that Glycyrrhiza uralensis acted against UC by activating the signaling of P-gp through upregualting its expression. The upregulation of FGFR4, SHP, and P-gp in liver conferred protective function in UC. CONCLUSION: Glycyrrhiza uralensis could regulate apoptosis of intestinal mucosal cells, through regulating the expression of apoptosis-related proteins and protective proteins of intestinal mucosa. The results provide novel options for UC treatment, as well as a rationale for pharmacology of Chinese traditional medicine, that is favorable for use of herbal medicine.

[Application of "process management and evaluation" in resident standardization training plan in acupuncture-moxibustion department of hospital for postgraduates of non-acupuncture- moxibustion speciality].

The teaching effect of "process management and evaluation" was assessed in resident standardization training plan in acupuncture-moxibustion department of hospital for postgraduates of non-acupuncture-moxibustion speciality. A total of 120 postgraduates of non-acupuncture-moxibustion speciality participating in resident standardization training were randomized into an observation group (60 cases) and a control group (60 cases, 1 case dropped off). In the control group, the conventional training mode was used. In the observation group, the "process management and evaluation" was adopted, in which, the syllabus was refined, various teaching modes were cooperated and the summary was conducted once a week. The training results were evaluated at the end of 1-month shift test and questionnaire was issued in all of the postgraduates of the two groups. In the observation group, the score for theory and the score of each of the items for technical ability, named differentiation and treatment, technical manipulation and physician-patient communication, as well as the total score were all higher than the control group successively (P<0.05, P<0.01). The results of the student questionnaire showed that in the items as "being liable to the memory of relevant knowledge" "connection of theory with practical ability" "stimulating students' interest and subjective initiative" "self-learning ability" "clinical question handling ability" and "communication ability with patients" as well as the total score in the observation group were all higher than the control group successively (P<0.01, P<0.05). The teaching effect of "process management and evaluation" is obviously better than the conventional teaching mode.

Simultaneous determination of nine constituents by validated UFLC-MS/MS in the plasma of cough variant asthma rats and its application to pharmacokinetic study after oral administration of Huanglong cough oral liquid.

Huanglong cough oral liquid (HL), an important traditional Chinese medicine prescription for treating pediatric cough variant asthma (CVA) in Nanjing hospital of traditional Chinese medicine for many years. In this study, a selective, accurate and sensitive ultra fast liquid chromatography extreme resolution coupled with mass spectrometer (UFLC-MS/MS) method was established and validated for the simultaneous determination of nine constituents including morusin, ephedrine, praeruptorin A, praeruptorin B, luteolin, rosmarinic acid, quercetin, amygdalin, caffeic acid in CVA rat plasma sensitized and challenged with ovalbumin and cinnamaldehyde. Plasma samples were prepared by protein precipitation with four-fold amount of methanol. UFLC separation was performed on a Thermo Scientific AcclaimTM RSLC 120 C18 column (2.1 mm × 100 mm, 2.2 µm) with mobile phase containing methanol and 0.1% formic acid-water by gradient elution in 8.1 min at total flow of 0.3 mL/min. The determination of target compounds in plasma was operated by multiple reaction monitoring (MRM) mode with positive and negative electrospray ionization (ESI) source. The correlation coefficients (r) of all compounds were from 0.9930 to 0.9994 in the linear range. Lower limit of quantification (LLOQ, ng/mL) was 0.81, 2.01, 2.11, 1.17, 1.04, 0.89, 0.67, 1.45 and 0.59 for morusin, ephedrine, praeruptorin A, praeruptorin B, luteolin, rosmarinic acid, quercetin, amygdalin and caffeic acid, respectively. Intra- and inter-day accuracy and precision, extraction recovery, matrix effect, carryover effect, dilution integrity, and stability were within the limits specified. The established method was effectively applied to a pharmacokinetic study of the nine compounds in CVA rat plasma following oral administration HL exact (7.5, 15, 30 g/kg).

Curcumin Regulates ERCC1 Expression and Enhances Oxaliplatin Sensitivity in Resistant Colorectal Cancer Cells through Its Effects on miR-409-3p.

BACKGROUND: Oxaliplatin (L-OHP) resistance is a major obstacle to the effective treatment of colorectal cancer. The resistance mechanism(s) of colorectal tumors to L-OHP may be related to the regulation of ERCC1 by cancer-expressed miRNAs, but no in-depth studies on the miRNAs that affect drug resistance have been performed. Curcumin (Cur) can reverse the drug resistance of cancer cells, but its effects on ERCC1 expression and miRNA profiles in colorectal cancer have not been studied. METHODS: To study the regulation effect of curcumin on ERCC1 expression and its effects on miRNAs, the L-OHP-resistant colorectal cancer cell line HCT116/L-OHP was established. MTT assays were used to evaluate cell proliferation. Flow cytometry was used to investigate apoptotic induction. Western blot and RT-PCR analysis were used to evaluate the expression of drug-associated ERCC1, Bcl-2, GST-π, MRP, P-gp, and survivin. RESULTS: HCT116//L-OHP cell lines were successfully established. The combination of L-OHP and curcumin could reduce L-OHP resistance in vitro. In addition, combination therapy inhibited the expression of ERCC1, Bcl-2, GST-π, MRP, P-gp, and survivin at the mRNA and protein level. Curcumin was found to inhibit ERCC1 through its ability to modulate miR-409-3p. CONCLUSION: Curcumin can overcome L-OHP resistance in colorectal cancer cells through its effects on miR-409-3p mediated ERCC1 expression.

Epigallocatechin gallate inhibits dimethylhydrazine-induced colorectal cancer in rats.

BACKGROUND: Epigallocatechin gallate (EGCG) is a polyhydroxy phenolic compound extracted from tea and its antitumor effect has received widespread attention. We explored the inhibitory effect of EGCG on dimethylhydrazine (DMH)-induced colorectal cancer (CRC) using a rat model, predicted the interaction between EGCG and CRC target genes using a database, and explained the EGCG associated target pathways and mechanisms in CRC. AIM: To understand the inhibitory mechanisms of EGCG on CRC cell proliferation and identify its pharmacological targets by network pharmacology analysis. METHODS: DMH (40 mg/kg, s.c., twice weekly for eight weeks) was used to induce CRC in rats. After model establishment, the rats were administered with EGCG (50, 100, or 200 mg/kg, p.o., once daily for eight weeks) and killed 12 and 20 wk after the start of the experiment. Formation of aberrant crypt foci and tumor was studied by histological analysis. Using network pharmacology analysis, candidate and collective targets of EGCG and CRC were identified, and Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes analyses were used to predict the pathways altered by EGCG. RESULTS: At week 12, high-dose EGCG treatment significantly reduced the tumor formation rate, total number of tumors, cancerous and non-cancerous tumors, tumor volume, ascites formation, and aberrant crypt foci count. At week 20, all three doses of EGCG were effective. Seventy-eight collective targets of EGCG and CRC were identified, of which 28 genes were dysregulated in CRC. Kyoto Encyclopedia of Genes and Genomes and GO analyses showed that the dysregulated genes were enriched in hsa05210 (CRC), hsa04115 (p53 signaling pathway), and hsa04151 (PI3K-Akt signaling pathway), GO:0043124 (negative regulation of I-kappaB kinase/NF-kappaB signaling pathway), GO:0043409 (negative regulation of mitogen-activated protein kinase cascade), and GO:2001244 (positive regulation of intrinsic apoptotic signaling pathway) respectively. CONCLUSION: EGCG inhibits the formation of DMH-induced CRC by regulating key pathways involved in tumorigenesis.

Dihydroartemisinin alleviates bile duct ligation-induced liver fibrosis and hepatic stellate cell activation by interfering with the PDGF-ßR/ERK signaling pathway.

Liver fibrosis represents a frequent event following chronic insult to trigger wound healing responses in the liver. Activation of hepatic stellate cells (HSCs), which is a pivotal event during liver fibrogenesis, is accompanied by enhanced expressions of a series of marker proteins and pro-fibrogenic signaling molecules. Artemisinin, a powerful antimalarial medicine, is extracted from the Chinese herb Artemisia annua L., and can inhibit the proliferation of cancer cells. Dihydroartemisinin (DHA), the major active metabolite of artemisinin, is able to attenuate lung injury and fibrosis. However, the effect of DHA on liver fibrosis remains unclear. The aim of this study was to investigate the effect of DHA on bile duct ligation-induced injury and fibrosis in rats. DHA improved the liver histological architecture and attenuated collagen deposition in the fibrotic rat liver. Experiments in vitro showed that DHA inhibited the proliferation of HSCs and arrested the cell cycle at the S checkpoint by altering several cell-cycle regulatory proteins. Moreover, DHA reduced the protein expressions of a-SMA, α1 (I) collagen and fibronectin, being associated with interference of the platelet-derived growth factor ß receptor (PDGF-ßR)-mediated ERK pathway. These data collectively revealed that DHA relieved liver fibrosis possibly by targeting HSCs via the PDGF-ßR/ERK pathway. DHA may be a therapeutic antifibrotic agent for the treatment of hepatic fibrosis.

Diallyl trisulfide attenuates ethanol-induced hepatic steatosis by inhibiting oxidative stress and apoptosis.

Inhibiting the major characteristics of alcoholic fatty liver (AFL) such as lipid accumulation, oxidative stress and apoptosis is a promising strategy of treating AFL. Diallyl trisulfide (DATS) is the major constituent isolated from garlic, which shows promise in the treatment of chronic liver disease. However, the effects of DATS on ethanol-induced liver injury and the related mechanisms remain unclear. The aim of this study was to evaluate the potential protective effects of DATS on AFL and the potential mechanisms. A single intragastric dose of ethanol was given to rats in vivo, while ethanol-stimulated LO2 cells were used as an in vitro model. Our results demonstrated that DATS prevented ethanol-induced injury, as indicated by the reduced activities of aspartate transaminase (AST) and alanine aminotransferase (ALT) in the serum and culture medium, and inhibition of cell apoptosis. Furthermore, DATS reduced hepatic steatosis by up-regulating the expression of peroxisome proliferator-activated receptor-alpha (PPAR-α) and down-regulating the expression of sterolregulatory element binding protein 1c(SREBP-1c). In addition, DATS alleviated ethanol-induced oxidative stress by enhancing non-enzymatic antioxidant and enzymatic antioxidants contents and by reducing the levels of reactive oxygen species (ROS) and malondialdehyde (MDA). These data collectively revealed that DATS protected ethanol-induced liver injury by inhibiting lipid accumulation and oxidative stress.

Diallyl trisulfide protects against ethanol-induced oxidative stress and apoptosis via a hydrogen sulfide-mediated mechanism.

Garlic is one natural source of organic sulfur containing compounds and has shown promise in the treatment of chronic liver disease. Dietary garlic consumption is inversely correlated with the progression of alcoholic fatty liver (AFL), although the exact underlying mechanisms are not clear. Our previous studies also have shown that diallyl trisulfide (DATS), the primary organosulfur compound from Allium sativum L, displayed anti-lipid deposition and antioxidant properties in AFL. The aim of the present study was to clarify the underlying mechanisms. In the present study, we used the intragastric infusion model of alcohol administration and human normal liver cell line LO2 cultured with suitable ethanol to mimic the pathological condition of AFL. We showed that accumulation of intracellular reactive oxygen species (ROS) was lowered significantly by the administration of DATS, but antioxidant capacity was increased by DATS. Additionally, DATS inhibited hepatocyte apoptosis via down-regulating Bax expression and up-regulating Bcl-2 expression, and attenuated alcohol-induced caspase-dependent apoptosis. More importantly, using iodoacetamide (IAM) to block hydrogen sulfide (H2S) production from DATS, we noted that IAM abolished all the above effects of DATS in ethanol-treated LO2 cells. Lastly, we found DATS could increase the expressions of cystathionine gamma-lyase (CSE) and cystathionine beta-synthase (CBS), the major H2S-producing enzymes. These results demonstrate that DATS protect against alcohol-induced fatty liver via a H2S-mediated mechanism. Therefore, targeting H2S may play a therapeutic role for AFL.

Dihydroartemisinin prevents liver fibrosis in bile duct ligated rats by inducing hepatic stellate cell apoptosis through modulating the PI3K/Akt pathway.

As a frequent event following chronic insult, liver fibrosis triggers wound healing reactions, with extracellular matrix components accumulated in the liver. During liver fibrogenesis, activation of hepatic stellate cells (HSCs) is the pivotal event. Fibrosis regression can feasibly be treated through pharmacological induction of HSC apoptosis. Herein we showed that dihydroartemisinin (DHA) improved liver histological architecture, decreased hepatic enzyme levels, and inhibited HSCs activation in the fibrotic rat liver. DHA also induced apoptosis of HSCs in such liver, as demonstrated by reduced distribution of α-SMA-positive cells and the presence of high number of cleaved-caspase-3-positive cells in vivo, as well as by down-regulation of Bcl-2 and up-regulation of Bax. In addition, in vitro experiments showed that DHA significantly inhibited HSC proliferation and led to dramatic morphological alterations in HSCs. we found that DHA disrupted mitochondrial functions and led to activation of caspase cascades in HSCs. Mechanistic investigations revealed that DHA induced HSC apoptosis through disrupting the phosphoinositide 3-kinase (PI3K)/Akt pathway and that PI3K specific inhibitor LY294002 mimicked the pro-apoptotic effect of DHA. DHA is a promising candidate for the prevention and treatment of liver fibrosis.

Herbal SGR Formula Prevents Acute Ethanol-Induced Liver Steatosis via Inhibition of Lipogenesis and Enhancement Fatty Acid Oxidation in Mice.

Our previous study indicated that herbal SGR formula partially attenuates ethanol-induced fatty liver, but the underlying mechanisms remain unclear. In the present study, mice were pretreated with SGR (100 and 200 mg/kg/d bw) for 30 d before being exposed to ethanol (4.8 g/kg bw). The biochemical indices and histopathological changes were examined to evaluate the protective effects and to explore potential mechanisms by investigating the adiponectin, tumor necrosis factor-α (TNF-α), peroxisome proliferators-activated receptor-α (PPAR-α), sterol regulatory element binding protein-1c (SREBP-1c), adenosine monophosphate-activated protein kinase (AMPK), and so forth. Results showed that SGR pretreatment markedly inhibited acute ethanol-induced liver steatosis, significantly reduced serum and hepatic triglyceride (TG) level, and improved classic histopathological changes. SGR suppressed the protein expression of hepatic SREBP-1c and TNF-α and increased adiponectin, PPAR-α, and AMPK phosphorylation in the liver. Meanwhile, acute toxicity tests showed that no death or toxic side effects within 14 days were observed upon oral administration of the extracts at a dose of 16 g/kg body wt. These results demonstrate that SGR could protect against acute alcohol-induced liver steatosis without any toxic side effects. Therefore, our studies provide novel molecular insights into the hepatoprotective effect of SGR formula, which may be exploited as a therapeutic agent for ethanol-induced hepatosteatosis.

Diallyl trisulfide attenuates carbon tetrachloride-caused liver injury and fibrogenesis and reduces hepatic oxidative stress in rats.

Liver fibrosis represents a frequent event following chronic insult to trigger wound healing reactions with accumulation of extracellular matrix (ECM) components in the liver. Activation of hepatic stellate cells (HSCs) is the pivotal event during liver fibrogenesis. The process of HSC activation is accompanied by enhanced expression of a series of marker proteins and pro-fibrogenic signal molecules. Natural products have been an important source of antifibrotic remedies. The present study aims to evaluate the in vivo effects of diallyl trisulfide (DATS), the primary component derived from garlic, on carbon tetrachloride (CCl4)-induced injury and fibrosis in rats. Our results showed that DATS improved liver histological architecture and decreased hepatic enzyme levels, but did not significantly affect cytochrome P450 2E1 activity in vivo. DATS also attenuated collagen deposition and inhibited HSC activation in the rat fibrotic liver demonstrated by reduced expression of α-smooth muscle actin, α1(I) procollagen, and fibronectin-three key markers of HSC activation-and by downregulation of transforming growth factor-ß receptor 1, platelet-derived growth factor-ß receptor, and epidermal growth factor receptor-three key receptors transmitting pro-fibrogenic pathways. In addition, DATS ameliorated hepatic oxidative stress by diminishing the levels of lipid peroxides and malondialdehyde and enhancing glutathione content. These data collectively revealed that DATS protected the rat liver from CCl4-caused injury and fibrogenesis in vivo, which was associated with inhibition of HSC activation and attenuation of oxidative stress. Our results suggested DATS as a promising antifibrogenic candidate for the prevention and treatment of liver fibrosis.

Paeonol inhibits hepatic fibrogenesis via disrupting nuclear factor-κB pathway in activated stellate cells: in vivo and in vitro studies.

BACKGROUND AND AIMS: Hepatic fibrosis represents a major cause of morbidity and mortality worldwide. The present study was to evaluate the antifibrogenesis effect of paeonol and involved mechanisms. METHODS: The degree of liver injury was evaluated biochemically by measuring serum and fibrotic markers and pathological examination. Cell viability was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and trypan blue staining. Cytotoxic effects were determined using lactate dehydrogenase release assay. Cell cycle was determined using single dyeing methods of propidium iodide (PI) by flow cytometry. Apoptosis was confirmed using double-staining of annexin V/PI and Hoechst. Western blot, immunofluorescence and real-time polymerase chain reaction were used to explore the molecular mechanisms. RESULTS: Treatment with paeonol significantly protected the liver from injury by reducing the activities of serum aspartate aminotransferase, alanine aminotransferase, improving the histological architecture of the liver, and by inhibiting activation of hepatic stellate cells (HSCs) in vivo. Interestingly, paeonol had no apparent cytotoxic effects but could markedly inhibit primary HSC proliferation and induced HSC cell cycle arrest at the G2/M checkpoint. These effects were caused by paeonol suppression of phosphorylation of cycle protein cdc2 and of CDK2. Moreover, that paeonol triggered mitochondrial apoptosis pathway and led to activation of caspase cascades in HSCs was found. Mechanistic investigations revealed that the nuclear factor-κB (NF-κB) pathway inhibition resulted in the earlier events. Furthermore, paeonol altered the expression of some marker proteins relevant to HSCs activation. CONCLUSION: Paeonol could inhibit HSC proliferation and induce mitochondrial apoptosis via disrupting NF-κB pathway, which might be the mechanisms of paeonol reduction of liver fibrosis.

Acupuncture combined with curcumin disrupts platelet-derived growth factor ß receptor/extracellular signal-regulated kinase signalling and stimulates extracellular matrix degradation in carbon tetrachloride-induced hepatic fibrosis in rats.

BACKGROUND: Acupuncture treatment has been increasingly used to treat chronic liver diseases. We previously reported that acupuncture combined with curcumin, a natural antifibrotic compound, could remarkably attenuate liver fibrosis in chemically intoxicated rats, but the underlying molecular mechanisms are poorly understood. The present study was aimed at investigating the effects of acupuncture combined with curcumin on platelet-derived growth factor (PDGF) signalling and extracellular matrix (ECM) regulation in the fibrotic liver. METHODS: A total of 60 Sprague-Dawley male rats were randomly divided into control, model, sham, acupuncture, curcumin and combination treatment groups. During the establishment of fibrosis using carbon tetrachloride (CCl(4)), acupuncture at LR3, LR14, BL18 and ST36 and/or curcumin treatment by mouth were performed simultaneously. After treatment, serum PDGF levels were measured. Protein and mRNA expression of key effectors in PDGF pathway and fibrinolysis in the liver was determined. RESULTS: Acupuncture combined with curcumin potently reduced serum PDGF levels and selectively disrupted the PDGF-ßR/extracellular signal-regulated kinase (ERK) cascade. Combination treatment also significantly repressed expression of connective tissue growth factor and upregulated expression of matrix metalloproteinase-9, promoting fibrinolysis in the fibrotic liver. CONCLUSIONS: The beneficial effects of acupuncture and its combination with curcumin could be attributed to the disruption of PDGF-ßR/ERK pathway and stimulated ECM degradation in the fibrotic liver. Acupuncture treatment significantly enhanced curcumin effects at the molecular level. These findings may provide molecular insights into the potential of acupuncture combined with curcumin for prevention of hepatic fibrosis.

[Effects of acupuncture intervention on hepatic platelet-derived growth factor signaling pathway in CCl4-induced hepatic fibrosis rats].

OBJECTIVE: To observe the effect of acupuncture stimulation of "Taichong" (LR 3), "Qimen" (LR 14), etc. on hepatic platelet-derived growth factor (PDGF) signal pathway activity at the protein and mRNA levels in hepatic fibrosis rats. METHODS: Forty-six SD rats were randomly divided into control (10 rats), model (12 rats), acupuncture (12 rats) and non-acupoint (12 rats) groups. Hepatic fibrosis model was established by intraperitoneal injection of mixture solution of 50% CCl4 and olive oil [1:1, 3 times on the 1st week (W), twice/W thereafter for 5 more weeks]. During modeling, acupuncture stimulation of "Taichong" (LR 3), "Qimen" (LR 14), "Ganshu" (BL 18) and "Zusanli" (ST 36) was conducted simultaneously. At the end of the experiments, all the rats were sacrificed for collecting their liver and blood samples, followed by separation of the hepatic stellate cells (HSCs). ELISA, Western blot and Real-time quantitative PCR techniques were used to detect the content of serum PDGF and expression levels of PDGF-beta receptor (PDGF-beta R), extracellular signal-regulated kinase (ERK1/2), c-jun N-terminal kinase (JNK) and P 38 genes and proteins of HSCs, respectively. RESULTS: Compared to the control group, serum PDGF content, and expression levels of PDGF-beta R mRNA and protein, ERK mRNA and protein and P 38 protein of HSCs in the model group were upregulated significantly (P < 0.01, P < 0.05). In comparison with the model group, serum PDGF content, and the expression levels of PDGF-beta R mRNA and protein, ERK mRNA and protein of HSCs in the acupuncture group were down-regulated apparently (P < 0.05, P < 0.01). No significant differences were found between the acupuncture and non-acupoint groups in serum PDGF content and between the model group and non-acupoint group in the expression levels of PDGF-beta R mRNA and protein, ERK mRNA and protein, JNK protein and P 38 protein of HSCs, as well as between the model group and acupuncture group in the expression levels of JNK protein and P 38 protein of HSCs (P > 0.05). CONCLUSION: Acupuncture intervention can effectively down-regulate serum PDGF content, and expression levels of PDGF-beta R mRNA and protein, ERK mRNA and protein of HSCs in liver fibrosis rats, which may contribute to its effect in improving liver fibrosis through down-regulating PDGF signal pathway activity.

Acupuncture combined with curcumin attenuates carbon tetrachloride-induced hepatic fibrosis in rats.

BACKGROUND: Increasingly, studies demonstrate the effectiveness of acupuncture therapy against liver fibrosis. Curcumin is a natural product with antifibrotic effects, but has poor pharmacokinetic profiles. This study aimed to evaluate whether acupuncture combined with curcumin could more potently attenuate liver fibrosis in chemical intoxicated rats. METHODS: 60 Sprague-Dawley male rats were randomly divided into control, model, sham, acupuncture, curcumin and combination therapy groups. During the establishment of fibrosis using carbon tetrachloride (CCl(4)), acupuncture at LR3, LR14, BL18 and ST36 and/or curcumin treatment by mouth were performed simultaneously. After treatment, pathological indexes and histology for hepatic injury and fibrogenesis were detected. The expression of extracellular matrix (ECM) components was also determined. RESULTS: Acupuncture combined with curcumin potently protected the liver from CCl(4)-induced injury and fibrogenesis, as indicated by reduced levels of serum aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, hyaluronic acid, laminin and procollagen III. Combined use also led to significant liver histological improvements. Furthermore, combined use effectively inhibited ECM expression such as α-smooth muscle actin, fibronectin and α1(1) collagen. CONCLUSIONS: Acupuncture treatment could significantly enhance the antifibrotic efficacy of curcumin on CCl(4)-induced hepatic fibrosis in rats in vivo, suggesting that a combination of acupuncture with curcumin may be exploited for the prevention of hepatic fibrosis.