Anemia and porphyria caused by N,N′-methylene-bis-acrylamide (MBA) in Mice and rats

The effects of N,N-methylene-bis-acrylamide (MBA), a cross-linking agent, on blood and bone marrow after repeated oral doses, were studied in mice and rats. Body weight, three major elements of the blood — erythrocytes, leucocytes and platelets — reticulocytes and bone marrow cells, were all reduced in either or both animals, especially in mice. Phenobarbital (PB) treatment did not greatly modify the effects of MBA in mice. An increase in free erythrocyte porphyrins and a decrease in ALA-D activity were observed in both animals. Urinary porphyrins were elevated in rats after MBA-dosing. PB-treatment did not significantly affect the elevation of porphyrins. After cessation of the MBA-dosing, all these changes were inclined to be restored to normal levels. Amounts of liver total porphyrins and microsomal P-450, and red cell fragility were within normal ranges in mice.     

Genistein alleviates pressure overload‐induced cardiac dysfunction and interstitial fibrosis in mice

Background and Purpose

Pressure overload‐induced cardiac interstitial fibrosis is viewed as a major cause of heart failure in patients with hypertension or aorta atherosclerosis. The purpose of this study was to investigate the effects and the underlying mechanisms of genistein, a natural phytoestrogen found in soy bean extract, on pressure overload‐induced cardiac fibrosis.

Experimental Approach

Genisten was administered to mice with pressure overload induced by transverse aortic constriction. Eight weeks later, its effects on cardiac dysfunction, hypertrophy and fibrosis were determined. Its effects on proliferation, collagen production and myofibroblast transformation of cardiac fibroblasts (CFs) and the signalling pathways were also assessed in vitro.

Key Results

Pressure overload‐induced cardiac dysfunction, hypertrophy and fibrosis were markedly attenuated by genistein. In cultured CFs, genistein inhibited TGFβ1‐induced proliferation, collagen production and myofibroblast transformation. Genistein suppressed TGFβ‐activated kinase 1 (TAK1) expression and produced anti‐fibrotic effects by blocking the TAK1/MKK4/JNK pathway. Further analysis indicated that it up‐regulated oestrogen‐dependent expression of metastasis‐associated gene 3 (MTA3), which was found to be a negative regulator of TAK1. Silencing MTA3 by siRNA, or inhibiting the activity of the MTA3‐NuRD complex with trichostatin A, abolished genistein''s anti‐fibrotic effects.

Conclusions and Implications

Genistein improved cardiac function and inhibited cardiac fibrosis in response to pressure overload. The underlying mechanism may involve regulation of the MTA3/TAK1/MKK4/JNK signalling pathway. Genistein may have potential as a novel agent for prevention and therapy of cardiac disorders associated with fibrosis.

Linked Articles

This article is part of a themed section on Chinese Innovation in Cardiovascular Drug Discovery. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-23

Abbreviations

CFs

cardiac fibroblasts

CTGF

connective tissue growth factor

ECM

extracellular matrix

EdU

5‐ethynyl‐2′‐deoxyuridine

EF

ejection fraction

ER

oestrogen receptors

FS

fractional shortening

MTA3

metastasis‐associated gene 3

NuRD

nucleosome remodelling and deacetylase

TAC

transverse aortic constriction

TAK1

TGFβ‐activated kinase 1

TSA

trichostatin A

Tables of Links

Curcumin attenuates cardiac fibrosis in spontaneously hypertensive rats through PPAR-γ activation

Aim:

To investigate the effects of curcumin (Cur) on cardiac fibrosis in spontaneously hypertensive rats (SHRs) and the mechanisms underlying the anti-fibrotic effect of Cur in rat cardiac fibroblasts (CFs) in vitro.

Methods:

SHRs were orally treated with Cur (100 mg·kg⁻¹·d⁻¹) or Cur (100 mg·kg⁻¹·d⁻¹) plus the PPAR-γ antagonist GW9662 (1 mg·kg⁻¹·d⁻¹) for 12 weeks. Cultured CFs were treated with angiotensin II (Ang II, 0.1 μmol/L) in vitro. The expression of relevant proteins and mRNAs was analyzed using Western blotting and real-time PCR, respectively. The expression and activity of peroxisome proliferator-activated receptor-γ (PPAR-γ) were detected using Western blotting and a DNA-binding assay, respectively.

Results:

Treatment of SHRs with Cur significantly decreased systolic blood pressure, blood Ang II concentration, heart weight/body weight ratio and left ventricle weight/body weight ratio, with concurrently decreased expression of connective tissue growth factor (CTGF), plasminogen activator inhibitor (PAI)-1, collagen III (Col III) and fibronectin (FN), and increased expression and activity of PPAR-γ in the left ventricle. Co-treatment with GW9662 partially abrogated the anti-fibrotic effects of Cur in SHRs. Pretreatment of CFs with Cur (5, 10, 20 μmol/L) dose-dependently inhibited Ang II-induced expression of CTGF, PAI-1, Col III and FN, and increased the expression and binding activity of PPAR-γ. Pretreatment with GW9662 partially reversed anti-fibrotic effects of Cur in vitro. Furthermore, pretreatment of CFs with Cur inhibited Ang II-induced expression of transforming growth factor-β1 (TGF-β1) and phosphorylation of Smad2/3, which were reversed by GW9662.

Conclusion:

Cur attenuates cardiac fibrosis in SHRs and inhibits Ang II-induced production of CTGF, PAI-1 and ECM in CFs in vitro. The crosstalk between PPAR-γ and TGF-β1/Smad2/3 signaling is involved in the anti-fibrotic and anti-proliferative effects of Cur.     

The NF-κB inhibitor celastrol attenuates acute hepatic dysfunction induced by cecal ligation and puncture in rats

Acute hepatic dysfunction associating sepsis is mediated mainly by toll-like receptor-4 (TLR-4)/nuclear factor kappa‐B (NF-κB) inflammatory pathway. This study explores potential hepatoprotective effect of the NF-κB inhibitor celastrol in cecal ligation and puncture (CLP) model in rats.Protective effect of celastrol (1 mg/kg, i.p., 1 h before CLP) was illustrated after 24 h by preventing CLP‐induced hepatic histopathological changes and elevation in serum hepatic biomarkers [alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TB) and gamma aminotransferase (γ-GT)] without affecting mortality. Celastrol anti‐inflammatory effect was illustrated by inhibiting increased serum and hepatic mRNA expression of interleukin-6 (IL-6) without affecting IL-10 elevation. Furthermore, celastrol inhibited CLP-induced elevations in hepatic mRNA expression of nuclear factor inhibitory protein kappa-B alpha (NFκBia), TLR-4, 5‐lipoxygenase (5-LOX) and prevented NF‐κB/p65 nuclear translocation and activation.In conclusion, celastrol prevented CLP-induced acute hepatic dysfunction through its anti-inflammatory effect by attenuating NF-κB activation, TLR-4 and 5-LOX expression with subsequent reduction in pro-inflammatory IL-6.     

Gender-related difference in the toxicity of 2-(2′-hydroxy-3′,5′-di-tert-butylphenyl)benzotriazole in rats: Relationship to the plasma concentration,in vitro hepatic metabolism,and effects on hepatic metabolizing enzyme activity

Previously, we showed that the toxic susceptibility of male rats to an ultraviolet absorber, 2-(2′-hydroxy- 3′,5′-di-tert-butylphenyl)benzotriazole (HDBB), was nearly 25 times higher than that of females. The present study aimed to clarify the mechanism of gender-related differences in HDBB toxicity. Male and female rats were given HDBB by gavage at 0.5, 2.5, or 12.5?mg/kg/day for 28 days, and plasma HDBB levels were measured at various time points by using liquid chromatography–tandem mass spectrometry. HDBB was rapidly absorbed and eliminated from the plasma in both sexes, and no sexual variations were found in the plasma levels. In the plasma, HDBB metabolites were not detected at any dose by the liquid chromatography–photodiode array detector. In an in vitro metabolic study using hepatic microsomes from male and female rats, HDBB was slightly metabolized, but no sexual differences were found in the residual HDBB ratio after a 60-minute incubation with an NADPH-generation system. Following 28-day HDBB administration, sexually different changes were found in cytochrome P450–dependent microsomal mixed-function oxidase activities in the liver. In males, 7-ethoxyresorufin O-deethylase activity decreased and lauric acid 12-hydroxylase activity increased at all doses. Decreases in aminopyrine N-demethylase activity and testosterone 2α- and 16α-hydroxylase activity were also found at 2.5?mg/kg and above in males. In females, the only significant change was increased lauric acid 12-hydroxylase activity at 12.5?mg/kg. These findings indicate that HDBB would have hepatic peroxisome proliferative activity, and the difference in susceptibility of male and female rats to this effect might lead to marked gender-related differences in HDBB toxicity.     

The effect of 3,4,3′,4′-tetrachlorobiphenyl on plasma retinol and hepatic retinyl palmitate hydrolase activity in female Sprague-Dawley rats

A single ip dose of 1, 5, or 15 mg/kg 3,4,3′,4′-tetrachlorobiphenyl (TCB) caused a dose-dependent depression of plasma retinol levels 24 hr after treatment of female Sprague-Dawley rats. The loss of plasma retinol appeared to be a function of depressed levels of the retinol-retinol-binding protein (RBP)-transthyretin ternary complex. No free retinol-RBP was observed in plasma from treated animals. Hepatic retinyl palmitate hydrolase (RPH) activity was also depressed and highly and positively correlated to the plasma retinol levels. TCB was determined to be a noncompetitive inhibitor of partially purified RPH with a K_I of 91 μm. Incubation of TCB with liver microsomes and NADPH decreased the inhibition of RPH. Doses of either 2,4,5,2′,4′,5′-hexachlorobiphenyl (HCB) or 3,4,5,3′,4′,5′-HCB equimolar to the 15 mg/kg TCB dose failed to cause a similar depression of plasma retinol in treated female rats. We conclude that, unlike other polychlorinated biphenyl congeners, TCB causes a depression of plasma retinol by inhibition of hepatic RPH.     

Inhibition of TNF-α protects against hepatic ischemia–reperfusion injury in rats via NF-κB dependent pathway

Hepatic ischemia–reperfusion injury (I/R) is a serious health problem associated with liver transplantation, resection surgery, and various types of shock especially hemorrhagic shock. In the present investigation, the effect of inhibition of tumor necrosis factor-alpha (TNF-α) using pentoxifylline or infliximab against hepatic I/R injury induced in rats by 45-min ischemia and 1-h reperfusion was studied. It was observed that both pentoxifylline and infliximab-treated groups showed a significantly lower extent and severity of liver injury. This is attributed to (1) a decrease in oxidative stress markers, (2) reduction of the expression of TNF-α, TNF-α type-1 receptors, and nuclear factor kappa B (NF-κB). Thus TNF-α inhibition may be one of the therapeutic interventions to overcome the deleterious effects of I/R on liver via reduction of oxidative stress and inhibition of inflammatory cascade.     

Kaempferol regulates the lipid-profile in high-fat diet-fed rats through an increase in hepatic PPARα levels

The aim of this study was to investigate the antiobesity and antihyperlipidemic effects of the flavonoid kaempferol (3,5,7,4'-tetrahydroxyflavone). After being fed a high-fat diet (HFD) for two weeks, rats were dosed orally with kaempferol (75, 150, or 300 mg/kg) or fenofibrate (100?mg/kg) once daily for eight weeks. Fenofibrate is an antilipemic agent that exerts its therapeutic effects through activation of peroxisome proliferator-activated receptor α (PPAR α). Kaempferol (300?mg/kg/day) produced effects similar to fenofibrate in reducing body weight gain, visceral fat-pad weights, plasma lipid levels, as well as the coronary artery risk and atherogenic indices of HFD-fed rats. Kaempferol also caused dose-related reductions in hepatic triglyceride and cholesterol content and lowered hepatic lipid droplet accumulation and the size of epididymal adipocytes in HFD-fed rats. Kaempferol and fenofibrate reversed the HFD-induced downregulation of hepatic PPAR α. HFD-induced reductions in the hepatic levels of acyl-CoA oxidase (ACO), and cytochrome P450 isoform 4A1 (CYP4A1) proteins were reversed by kaempferol and fenofibrate. The elevated expression of hepatic sterol regulatory element binding proteins (SREBPs) in HFD-fed rats were lowered by kaempferol and fenofibrate. These results suggest that kaempferol reduced the accumulation of visceral fat and improved hyperlipidemia in HFD-fed obese rats by increasing lipid metabolism through the downregulation of SREBPs and promoting the hepatic expression of ACO and CYP4A1, secondary to a direct upregulation hepatic PPAR α expression.     

Evodiamine ameliorates liver fibrosis in rats via TGF-β1/Smad signaling pathway

Liver fibrosis is considered to be a result of chronic liver pathological changes, and hepatic stellate cells (HSCs) play an important role during this process. Evodiamine, an indole alkaloid derived from Evodia rutaecarpa, exhibits pharmacological activities. This study focused on the effects of evodiamine on carbon tetrachloride (CCl₄)-induced liver fibrosis in rats and HSCs in vitro via the TGF-β1/Smad signaling pathway. A liver fibrosis rat model was established by the intraperitoneal injection of CCl₄ (3 ml/kg, 30% in olive oil). Evodiamine (15 and 25 mg/kg) was administered orally for 8 weeks. HSCs were treated with different evodiamine concentrations. The results indicated that evodiamine could improve the histopathological abnormalities in liver tissues and decrease the level of aspartate aminotransferase (AST), alanine aminotransferase (ALT), hydroxyproline, and total bilirubin (TBIL). Concentrations of IL-6, tumor necrosis factor-α (TNF-α), collagen-I (COL-I), and collagen-III (COL-III) were reduced by evodiamine. Western blotting and real-time PCR showed that protein expression of transforming growth factor-β (TGF-β1), p-Smad 2/3 (phosphorylation of Smad 2/3), and smooth muscle alpha-actin (α-SMA) as well as mRNA expression of TGF-β1 and α-SMA in liver tissues were downregulated by evodiamine. The cell proliferation, production of hydroxyproline, and the protein expression of TGF-β1, p-Smad 2/3, and α-SMA in HSCs were dose-dependently reduced by evodiamine. Collectively, evodiamine had an antifibrosis effect in CCl₄-induced liver fibrosis, and reduced HSCs proliferation and collagen metabolism in vitro. The major mechanism was downregulation of relative expression of TGF-β1, p-Smad 2/3, and α-SMA.     

Dihydroartemisinin alleviates bile duct ligation-induced liver fibrosis and hepatic stellate cell activation by interfering with the PDGF-βR/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 β receptor (PDGF-βR)-mediated ERK pathway. These data collectively revealed that DHA relieved liver fibrosis possibly by targeting HSCs via the PDGF-βR/ERK pathway. DHA may be a therapeutic antifibrotic agent for the treatment of hepatic fibrosis.     

PPARα-dependent modulation of hepatic CYP1A by clofibric acid in rats

Fibrates, hypolipidemic drugs, have been reported to suppress the metabolic activities of cytochrome P450 1A1 and 1A2 in rats but the mechanism has not been elucidated. In the present study we tested the hypothesis that the inhibitory effect of fibrates on arylhydrocarbon receptor (AhR) function may be due to their stimulatory effects on PPAR. Sudan III (S.III) treatment induced CYP 1A1 and CYP 1A2 protein expression, mRNA and their metabolic activities, methoxyresorufin-O-demethylase (MROD) and ethoxyresorufin-O-deethylase (EROD), in Wistar rats higher than those in the control. Co-treatment of rats with S.III and clofibric acid (CA) caused a 40–50% decrease in the induced levels of CYP1A1 and CYP1A2 protein, mRNA expression and their metabolic activities and reduced AhR protein expression. When we treated HepG2 cells with S.III and/or CA, no suppressive effect on S.III-induced CYP1A1 protein expression due to CA was found. HepG2 cells were transiently transfected with increasing concentrations of PPAR mammalian expression vector and exposed to the same treatment. CA co-treatment with S.III decreased AhR protein and S.III-induced CYP1A1 protein expression with increasing dose of PPAR transfected into HepG2 cells. Our results demonstrate that the suppressive effect of fibrates on CYP1A is PPAR-dependent and suggest that PPARhas an inhibitory effect on AhR function.Abbreviations AhR Arylhydrocarbon receptor - CA Clofibric acid - CYP1A1 Cytochrome P450 1A1 - CYP1A2 Cytochrome P450 1A2 - EROD Ethoxyresorufin-O-deethylase - MROD Methoxyresorufin-O-demethylase - PPAR Peroxisome proliferator activated receptor alpha - PPRE Peroxisome proliferator response element - RXR Retinoid-X receptor - S.III Sudan III - TCDD 2,3,7,8-tetrachlorodibenzo-p-dioxin - TNF Tumor necrosis factor-alpha - XAP2 Hepatitis B virus X-associated protein     

Parthenolide,an NF-κB inhibitor,alleviates peritoneal fibrosis by suppressing the TGF-β/Smad pathway

Transforming growth factor (TGF)-β/Smad signalling plays a central role in the pathogenesis of peritoneal fibrosis related to peritoneal dialysis (PD). Parthenolide (PTL), a naturally occurring phytochemical, is isolated from the shoots of feverfew (Tanacetum parthenium) and displays analgesia, anti-inflammation and anticancer activities. In this study, we examined the therapeutic potential of PTL on PD-related peritoneal fibrosis induced by daily intraperitoneal injection of 4.25% dextrose-containing PD fluid (PDF) in vivo and TGF-β1-induced epithelial-mesenchymal transition (EMT) in vitro. PTL was administered daily before PDF injection or after 14 days of PDF injection. Both PTL treatments showed a protective effect on peritoneal fibrosis and prevented peritoneal dysfunction. Similarly, PTL suppressed the expression of fibrotic markers (fibronectin and collagen I) and restored the expression of the epithelial marker (E-cadherin) in TGF-β1-treated HMrSV5 cells. Furthermore, PTL inhibited TGF-β1-induced Smad2 and Smad3 phosphorylation and nuclear translocation but did not influence Smad1/5/9 phosphorylation or activate other downstream signalling pathways of TGF-β1, including AKT, extracellular signal-regulated kinase (ERK) or p38. In conclusion, PTL treatment may represent an effective and novel therapy for PD-associated peritoneal fibrosis by suppressing the TGF-β/Smad pathway.     

β-Arrestin2 deficiency attenuates oxidative stress in mouse hepatic fibrosis through modulation of NOX4

Hepatic fibrosis is a disease characterized by excessive deposition of extracellular matrix(ECM)in the liver.Activation of hepatic stellate cells(HSCs)is respon...     

Genistein modulates NF-κB-associated renal inflammation, fibrosis and podocyte abnormalities in fructose-fed rats

The study determines the effect of genistein on inflammatory status and expression of nuclear factor-kappa B (NF-κB p65), transforming growth factor-β1 (TGF-β1) and receptor for advanced glycation end products (RAGE) in kidney of fructose-fed rats. Adult male Wistar rats were fed a diet containing either starch or fructose as the source of carbohydrate. Fifteen days later, after confirming the development of insulin resistance in fructose-fed rats, the rats in each dietary group were divided into two and treated with either genistein (1 mg/kg/day) in 30% dimethylsulfoxide (DMSO) or 30% DMSO alone for the next 45 days. The expression of NF-κB P(65), TGF-β1 and RAGE, histochemical localization of α-smooth muscle actin (α-SMA), levels of tumour necrosis factor-α (TNF-α) and interleukin-6(IL-6) and ultrastructural analysis were performed at the end of the experimental period. Fructose-fed rats displayed inflammatory changes in kidney. Increased expression of TGF-β1 and RAGE in cytosol and NF-κB p65 in nuclear fraction were observed. α-SMA expression was higher in fructose-fed rat kidney. Proliferation of connective tissue was evident from increased collagen deposition in perivascular and intraglomerular regions. Administration of genistein to fructose-fed rats reduced inflammation, fibrogenesis and NF-κB activation. Genistein also mitigated the structural changes such as basement membrane thickening, reduction in podocyte number and loss of glomerular filtration barrier integrity. These findings suggest that genistein prevents inflammation, fibrosis and early nephropathic changes in fructose-fed insulin resistant rats secondary to the attenuation of NF-κB activation.     

Über die Wirkung von Vitamin B12 und die Krebserzeugung durch Buttergelb

Zusammenfassung Der Verf. untersuchte die hepatische Gewebeatmung bei Ratten unter Behandlung mit Buttergelb und Vitamin B₁₂. Dabei wurde nachgewiesen, daß Vitamin B₁₂ die schon durch Buttergelb veränderte Atmungstätigkeit des präneoplastischen Lebergewebes noch bis zu 35% weiter progressiv senkt.Mit 1 Textabbildung     

The disposition of the new arabinosylcytosine derivative—5′-chloro-5′-deoxy-arabinosylcytosine—in rats

• 1.1. Pharmacokinetic properties of a new derivative of the widely used and very potent antileukemic agent arabinosylcytosine (araC)—5′-chloro-5′-deoxy-arabinosylcytosine (5′-Cl-araC)—were investigated after intraperitoneal (i.p.) and oral routes of administration in rats and compared with the equimolar dose of araC administered orally.
• 2.2. It was found that substitution of the hydroxyl group at position 5′ resulted in a change of pharmacokinetic parameters.
• 3.3. There is a large difference in average serum concentrations of 5′-Cl-araC administered by the i.p. and oral routes; the average serum concentration obtained after i.p. injection being several times higher in comparison to those after oral administration.
• 4.4. However, the latter are, at the same time, lower than the average serum concentrations of araC administered by the same route in an equimolar dose.
• 5.5. On the other hand, the apparent volume of distribution is much larger, and the area under the curve of serum concentration of 5′-Cl-araC is smaller, after oral as compared to the i.p. route of administration indicating more extensive tissue distribution together with higher tissue binding of 5′-Cl-araC when compared to the parental drug araC.

     

Puerarin alleviates vincristine-induced neuropathic pain and neuroinflammation via inhibition of nuclear factor-κB and activation of the TGF-β/Smad pathway in rats

Chemotherapy-induced neuropathic pain harms the quality of life patients. Vincristine is an often used chemotherapeutic drug that evokes neuralgia via inflammation. Puerarin (Pue) extracted from Puerariae Lobatae Radix has analgesic and anti-inflammatory effects; however, its possible effect and mechanism in vincristine (Vin)-induced neuropathic pain has not been investigated. The present research aimed to explore whether Pue could relieve chemotherapy-evoked neuropathic pain and the underlying mechanism actions. Rat neuropathic pain was established by intraperitoneal injection of vincristine. Pue was orally administered in two dose levels (25 or 50 mg/kg/d) for three weeks. The paw withdrawal latency and paw withdrawal threshold were performed to evaluate the pain behaviors. Inflammatory cytokines in the spinal cord and dorsal root ganglion were measured by ELISA kits. qRT-PCR, western blot, and immunofluorescence staining were employed to measure the level and expression feature of inflammatory cytokines. Our findings showed that Pue improved hyperalgesia and allodynia. Treatment with Pue restored the levels of tumor necrosis factor-α (TNF-α), and IL-1β and increased the levels of transforming growth factor-β (TGF-β), and interleukin-10 (IL-10). On the molecular level, treatment with Pue down-regulated the protein levels of IL-1β, and NF-κBp65 and up-regulated the protein levels of TGF-β, p-Smad2, and p-Smad3 (TGF-β/Smad) in the spinal cord and DRG. Immunofluorescence staining further demonstrated that Pue decreased the NF-κBp65 protein. Our findings imply that Pue relieved chemotherapy-induced neuropathic pain might be attributable to the suppression of inflammation cytokines. The anti-inflammation action of Pue might be associated with the activation of the TGF-β/Smad pathway, a novel mechanism exploring its prophylactic effect in vincristine-induced neuropathic pain.