Cuscuta chinensis Lam. Protects Against Light-Induced Retinal Degeneration: Therapeutic Implications for Photoreceptor Degenerative Disorders.

Cuscuta chinensis Lam. (CCL) is a medicinal herb widely used in traditional Chinese medicine for the treatment of ophthalmic diseases, including age-dependent vision-threatening retinal degenerative disorders that involve irreversible loss of the first-order retinal neurons, photoreceptors. However, evidence is lacking if CCL is pharmacologically active at protecting against loss of photoreceptors and photoreceptor degeneration-associated retinal structural and functional impairment. The current study thus evaluates the potential photoreceptor protective effects of CCL to better support its clinical applications in the prevention and treatment of photoreceptor degenerative diseases. Non-invasive full-retinal optical coherence tomography, electroretinography, histological examination, immunohistochemistry and real-time qPCR analysis were performed to assess the retinal protective effects of CCL in light-exposed BALB/c mice characterized by photooxidative stress-mediated photoreceptor loss and associated retinal morphological and functional impairment. The results showed that CCL treatment protected against light-induced degeneration of the photoreceptor structure and deterioration of the retinal function. Furthermore, CCL treatment increased the retinal expression of rhodopsin, S-opsin and M-opsin, supporting the protective effects of CCL in both rod and cone photoreceptors. CCL treatment suppressed photoreceptor cell death in the light-exposed retinas. The morphological integrity of the second-order retinal neurons was also preserved as a result of CCL treatment. In addition, CCL treatment attenuated light-induced reactive müller gliosis, microglial activation and inflammation in the retina. In conclusion, the current work demonstrates for the first time that CCL protects against photooxidative stress-mediated degeneration of photoreceptors and associated disturbance of structural, functional and immune homeostasis of the retina. The findings here thus provide novel experimental evidence supporting the clinical application of CCL in the prevention and treatment photoreceptor degenerative diseases.

Dual Activity of Ginsenoside Rb1 in Hypertrophic Cardiomyocytes and Activated Macrophages: Implications for the Therapeutic Intervention of Cardiac Hypertrophy.

PURPOSE: Owing to the important mechanistic implications in the pathogenesis of cardiac hypertrophy and heart failure, inflammation has been proposed as a druggable target for the treatment of cardiac hypertrophy and heart failure. Ginseng is a widely used medicinal herb for the treatment of cardiovascular disorders. As one of the major chemical components of ginseng, ginsenoside Rb1 (Rb1) contributes to the cardiovascular effects of ginseng. Meanwhile, anti-inflammatory activity of Rb1 has also been documented. The current work aims to further delineate the pharmacological implications of Rb1 in the treatment of cardiac hypertrophy. METHODS: Angiotensin II (Ang II) infusion mouse model was adopted to investigate the effects of Rb1 on cardiac hypertrophic remodeling and associated inflammation in vivo. Furthermore, the mechanisms of actions of Rb1 in modulating the hypertrophic and inflammatory responses were investigated in cardiomyocytes and macrophages, respectively. RESULTS: Rb1 mitigates Ang II-induced cardiac hypertrophy, cardiac inflammation and systemic inflammation in vivo. In cardiomyocytes, Rb1 directly counteracts the pro-hypertrophic effects of Ang II and phenylephrine and maintains the mitochondrial function. In lipopolysaccharide (LPS)-stimulated macrophages, Rb1 decreases the phosphorylation of mitogen-activated protein kinases (MAPKs) and mitogen-activated protein kinase kinase 1/2 (MEK1/2) and reduces the production of inflammation mediators such as interleukin (IL)-1 beta, IL-6 and tumor necrosis factor (TNF). Rb1 also suppresses the expression of pro-hypertrophic microRNA-155 (miR-155) in LPS- or Ang II-stimulated macrophages. Furthermore, in activated macrophages, miR-155 is in part accountable for the suppressive effect of Rb1 on the production of IL-6, an inflammation mediator with pro-hypertrophic functions in the heart. CONCLUSION: The work here provides novel experimental evidence supporting the notion that Rb1 protects against cardiac hypertrophy in part through suppressing the inflammatory mechanisms that promotes the pathological remodeling of the heart.

Deciphering the Effective Constituents and Mechanisms of Portulaca oleracea L. for Treating NASH via Integrating Bioinformatics Analysis and Experimental Pharmacology.

Nonalcoholic steatohepatitis (NASH) is a highly prevalent metabolic disorder. Currently, there are no effective pharmacotherapeutic options for preventing and treating NASH. Portulaca oleracea L. (POL) is an edible herb that has been used for preventing and treating some metabolic disorders in China, but the bioactive constituents in POL and the related mechanisms for treating NASH are still unclear. Here, a comprehensive research strategy was used to identify the core genes and the key constituents in POL for treating NASH, via integrating bioinformatics analysis and experimental pharmacology both in vitro and in vivo. The phenotypes and mechanisms of POL were carefully investigated by performing a set of in vivo and in vitro experiments. Bioinformatics analysis suggested that prostaglandin-endoperoxide synthase 2 (PTGS2) was the core target and myricetin (Myr) was the key constituent in POL for treating NASH. In NASH mice model induced by methionine choline deficiency diet, POL significantly alleviated hepatic steatosis and liver injury. In free fatty acids-induced hepatocytes, POL and Myr significantly down-regulated the expression of PTGS2, decreased the number of lipid droplets, and regulated the mRNA expression of lipid synthesis and homeostasis genes, including FASN, CPT1a, SERBP1c, ACC1, and SCD1. In lipopolysaccharide-induced macrophages, POL and Myr significantly reduced the expression of PTGS2 and blocked the secretion of inflammatory mediators TNF-α, IL-6, and IL-1ß. Further investigations demonstrate that Myr acts as both suppressor and inhibitor of PTGS2. Collectively, POL and its major component Myr can ameliorate NASH via down-regulating and inhibiting PTGS2, suggesting that POL and Myr can be developed as novel medicines for treating NASH.

Shihu Yeguang Pill protects against bright light-induced photoreceptor degeneration in part through suppressing photoreceptor apoptosis.

Photoreceptor cells are first-order retinal neurons that directly contribute to the formation of vision. Photoreceptor degeneration is the primary cause of vision impairment during the course of retinopathies such as retinitis pigmentosa and age-related macular degeneration, for which photoreceptor-targeted therapies are currently unavailable. Shihu Yeguang Pill (SYP), a classic formula in traditional Chinese medicine, has a long histology of clinical application for the treatment of a wide range of retinopathies in China. However, whether SYP is pharmacological effective at protecting photoreceptor cells is unclear. The current study thus directly addressed the pharmacological implications of SYP in photoreceptor degeneration in a mouse model characterized by bright light-induced retinal degeneration. Non-invasive full-retinal assessment was carried out to evaluate the effect of SYP on the retinal structure and function through optical coherence tomography and electroretinography, respectively. In addition, photoreceptor apoptosis, second-order neuron impairment and reactive changes in retinal microglial and müller cells, hallmark pathologies associated with photoreceptor degeneration, were assessed using immunohistochemistry and real-time PCR analyses. The results showed that SYP treatment attenuated bright light-induced impairment of the retinal structure and function. Moreover, SYP treatment suppressed photoreceptor apoptosis, alleviated the impairment of bipolar and horizontal cells and mitigated the reactive changes of müller and microglial cells in the bright light-exposed retinas. Real-time PCR analyses showed that dysregulated expression of pro-apoptotic c-fos and c-jun and anti-apoptotic bcl-2 as well as proinflammatory TNF-α in the bright light-exposed retinas was partially normalized as a result of SYP treatment. In summary, the work here demonstrates for the first time that SYP treatment protects the retinas from developing bright light-induced photoreceptor degeneration and associated alterations in second-order neurons and glial cells. The findings here thus provide experimental evidence to better support the mechanism-guided clinical application of SYP in the treatment of related retinal degenerative diseases.

Folic Acid Supplementation for Stroke Prevention in Patients With Cardiovascular Disease.

BACKGROUND: Controversy remains regarding the efficacy of folic acid supplementation in reducing the risk of stroke. This study aimed to evaluate the effect of folic acid supplementation on stroke prevention in patients with cardiovascular disease (CVD). MATERIALS AND METHODS: We searched the PubMed, EMBASE and Cochrane Library databases through October 2016 to identify randomized clinical trials of folic acid supplementation to prevent stroke in patients with CVD. Relative risks (RRs) with 95% CIs were used to examine the association between folic acid supplementation and the risk of stroke with a fixed-effect model. Stratified analyses were performed according to modifiers that may affect the efficacy of folic acid supplementation. RESULTS: Eleven studies with a total of 65,790 participants were included. Folic acid supplementation was associated with a significant benefit in reducing the risk of stroke in patients with CVD (RR = 0.90; 95% CI: 0.84-0.97; P = 0.005). In the stratified analysis, greater beneficial effects were observed in participants with a decrease in homocysteine concentrations of 25% or greater (RR = 0.85; 95% CI: 0.74-0.97; P = 0.03), those with a daily folate dose of less than 2mg (RR = 0.78; 95% CI: 0.68-0.89; P = 0.01), and populations in regions with no or partly fortified grain (RR = 0.87; 95% CI: 0.81-0.94; P = 0.04). CONCLUSIONS: Our meta-analysis demonstrated that folic acid supplementation is effective in stroke prevention in patients with CVD.

Apigenin-7-diglucuronide protects retinas against bright light-induced photoreceptor degeneration through the inhibition of retinal oxidative stress and inflammation.

Vision impairment in retinal degenerative diseases such as age-related macular degeneration is primarily associated with photoreceptor degeneration, in which oxidative stress and inflammatory responses are mechanistically involved as central players. Therapies with photoreceptor protective properties remain to be developed. Apigenin-7-diglucuronide (A7DG), a flavonoid glycoside, is present in an assortment of medicinal plants with anti-inflammatory or ant-oxidant activities. However, the pharmacological significance of A7DG remains unknown in vivo. The current study isolated A7DG from Glechoma longituba (Nakai) Kuprian and investigated the retinal protective effect A7DG in mice characterized by bright light-induced photoreceptor degeneration. The results showed that A7DG treatment led to remarkable photoreceptor protection in bright light-exposed BALB/c mice. Moreover, A7DG treatment alleviated photoreceptor apoptosis, mitigated oxidative stress, suppressed reactive gliosis and microglial activation and attenuated the expression of proinflammatory genes in bright light-exposed retinas. The results demonstrated for the first time remarkable photoreceptor protective activities of A7DG in vivo. Inhibition of bright light-induced retinal oxidative stress and retinal inflammatory responses was associated with the retinal protection conferred by A7DG. The work here warrants further evaluation of A7DG as a pharmacological candidate for the treatment of vision-threatening retinal degenerative disorders. Moreover, given the general implication of oxidative stress and inflammation in the pathogenesis of neurodegeneration, A7DG could be further tested for the treatment of other neurodegenerative disorders.

miR-29c is implicated in the cardioprotective activity of Panax notoginseng saponins against isoproterenol-induced myocardial fibrogenesis.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax notoginseng (Burkill) F.H. Chen (Araliaceae) has a long history of clinical application in China for the treatment of cardiovascular diseases. Panax notoginseng saponins (PNS) have been proven to be the major cardioprotective substances of Panax notoginseng (Burkill) F.H. Chen (Araliaceae). AIM OF THE STUDY: The current study further investigated the molecular mechanisms associated with the cardioprotective effect of PNS. MATERIALS AND METHODS: C57BL/6J mice were subject to isoproterenol (ISO)-induced myocardial injury in the absence or presence of PNS treatment. Histological, immunohistochemical and molecular biological approaches were taken to assess the effects of PNS treatment on ISO-induced myocardial injury and ensuing fibrogenesis. RESULTS: PNS treatment significantly attenuated ISO-induced myocardial injury and fibrosis. The expression of an anti-fibrotic microRNA, miR-29c, was significantly decreased in ISO-challenged mouse hearts. In contrast, PNS treatment resulted in increased cardiac expression of miR-29c. The expression of miR-29c target genes including Collagen (Col) 1a1, Col1a2, Col3a1 and Col5a1, fibrillin 1 (Fbn1) as well as TGFß1 was significantly increased by ISO, which exhibited decreased expression by PNS intervention. CONCLUSIONS: Our results demonstrate for the first time that the cardioprotective effects of PNS could in part implicate increased expression of miR-29c in the heart, which may help increase the understanding of the pharmacological activities of PNS in treating cardiovascular disorders.

Panax notoginseng saponins attenuate lung cancer growth in part through modulating the level of Met/miR-222 axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax notoginseng saponins (PNS) are the major chemical constituents of Panax notoginseng (Burkill) F.H. Chen (Araliaceae), a medicinal herb extensively used in China for the treatment of various diseases including cancer. PNS have been reported to contribute to the therapeutic effects of Panax notoginseng in disease conditions including lung cancer. AIM OF THE STUDY: The current study aims to further understand the molecular mechanisms implicated in the pharmacological activities of PNS in attenuating lung cancer growth. MATERIALS AND METHODS: Lewis lung carcinoma (LLC) cell line was employed and the impact of PNS treatment on the viability of LLC cells was first examined in vitro. The tumor-suppressive effect of PNS was further validated in vivo by assessing the tumor growth in BALB/c mice inoculated with LLC cells. Whole genome microarray and real-time PCR analyses were performed to examine and verify altered expression of genes associated with PNS treatment. Real-time PCR and western blotting analyses were also carried out to investigate the implication of microRNA (miRNA)-mediated gene expression regulation in the anti-tumor activity of PNS. RESULTS: PNS treatment resulted in selective impairment of the survival of LLC cells. Furthermore, PNS treatment led to attenuated growth of tumors derived from inoculated LLC cells in mice. Bioinformatic analyses of gene expression profiles revealed that multiple pathways associated with tumorigenesis were significantly modulated by PNS treatment in vivo. The expression of an array of genes promoting tumorigenesis and progression including Hgf, Met, Notch3, Scd1, Epas1, Col1a1, Raf1, Braf1 and CDK6 was significantly decreased by PNS treatment, whereas the expression of tumor suppressive Rxrg was significantly increased as a result of PNS treatment. The level of miR-222, a miRNA regulated by Met, was significantly decreased by PNS treatment. The expression of tumor suppressor p27 and PTEN, miR-222 target genes, was significantly increased by PNS treatment. CONCLUSION: Out work here presented novel evidence demonstrating that multiple mechanisms were implicated in the anti-tumor effects of PNS in lung cancer models. Particularly, PNS treatment significantly modulated the level of Met/miR-222 axis in LLC cells. Increased understanding of the anti-tumor mechanisms of PNS may provide further experimental evidence to help optimize the therapeutic modalities for the treatment of lung cancer and other types of cancer.

Yiqi Huaju formula, a Chinese herbal medicine, reduces arterial pressure in salt­sensitive hypertension by inhibiting renin­angiotensin system activation.

Hypertension is a chronic disease with a high prevalence, and is associated with a high risk of vascular disease and premature death. Traditional Chinese medicine has been administered to treat hypertension for many years. In the present study, the effects of Yiqi Huaju formula (YQ; a compound used in traditional Chinese herbal medicine) were observed in salt­sensitive hypertension, which was induced by a high­salt and high­fat (HSF) diet and the potential mechanism was investigated. YQ was prepared from five plant extracts and was dissolved in normal sodium chloride prior to use. Male Sprague­Dawley rats were randomly divided into three groups, and fed either a normal diet (control), an HSF diet or an HSF diet with YQ. At week eight, blood pressure was measured and 24­h urine samples were collected from all of the rats. The rats were subsequently sacrificed, and their blood was collected for biochemical analyses and kidney tissue samples were dissected for the immunohistochemical assay. YQ was observed to decrease the high arterial pressure and serum total cholesterol level, which had been induced by the HSF diet. It also enhanced the excretion of urinary angiotensinogen, Na+, and decreased the loss of urinary aldosterone, K+ and microalbuminuria. In addition, YQ inhibited the high mRNA expression level of renal renin, angiotensin II (Ang II), and Ang II receptor, type 1 (AT1R), and inhibited the protein expression of renal AT1R and Ang II receptor type 2, which had been induced by the HSF diet. These results indicate that YQ may reduce the arterial pressure in salt­sensitive hypertension via the inhibition of renin­angiotensin system activation.

17ß-estradiol prevents cardiac diastolic dysfunction by stimulating mitochondrial function: a preclinical study in a mouse model of a human hypertrophic cardiomyopathy mutation.

OBJECTIVE: We investigated the effect of ovariectomy (OVX) and 17ß-estradiol (E2) replacement on both mitochondrial and myocardial function in cTnT-Q92 transgenic mice generated by cardiac-restricted expression of a human hypertrophic cardiomyopathy (HCM) mutation. METHODS: The cTnT-Q92 mice were ovariectomized at twenty weeks of age and were treated with either placebo (OVX group) or E2 (OVX+E2 group) for twelve weeks before being sacrificed. Wild-type and cTnT-Q92 female mice receiving sham operation were used as controls. Indices of diastolic function such as mitral early (E) and late (A) inflow as well as isovolumic relaxation time (IVRT) were measured by echocardiography. A Clark-type electrode was used to detect respiratory control, and ATP levels were determined at the mitochondrial level using HPLC. Key components related to mitochondrial energy metabolism, such as peroxisome proliferator-activated receptor α (PPARα), PPARγ coactivator 1α (PGC-1α) and nuclear respiratory factor-1 (NRF-1), were also analyzed using Western blot and RT-PCR. The levels of oxidative stress markers were determined by measuring malondialdehyde (MDA) using the thiobarbituric acid assay. RESULTS: The cTnT-Q92 mice had impaired diastolic function compared with wild-type mice (E/A ratio, 1.39 ± 0.04 vs. 1.21 ± 0.01, p<0.001; IVRT, 19.17 ± 0.85 vs. 22.15 ± 1.43 ms, p=0.028). In response to ovariectomy, cardiac function further decreased compared with that observed in cTnT-Q92 mice that received the sham operation (E/A ratio, 1.15 ± 0.04 vs. 1.21 ± 0.01, p<0.001; IVRT, 28.31 ± 0.39 vs. 22.15 ± 1.43 ms, p=0.002). Myocardial energy metabolism, as determined by ATP levels (3.49 ± 0.31 vs. 5.07 ± 0.47 µmol/g, p<0.001), and the mitochondrial respiratory ratio (2.04 ± 0.10 vs. 2.63 ± 0.11, p=0.01) also decreased significantly. By contrast, myocardial concentrations of MDA increased significantly in the OVX group, and PGC-1α, PPARα and NRF-1decreased significantly. E2 supplementation significantly elevated myocardial ATP levels (4.55 ± 0.21 vs. 3.49 ± 0.31 µmol/g, p=0.003) and mitochondrial respiratory function (3.93 ± 0.05 vs. 2.63 ± 0.11, p=0.001); however, it reduced the MDA level (0.21 ± 0.02 vs. 0.36 ± 0.03 nmol/g, p<0.001), which subsequently improved diastolic function (E/A ratio, 1.35 ± 0.06 vs. 1.15 ± 0.04, p<0.001; IVRT, 18.22 ± 1.16 vs. 28.31 ± 0.39 ms, p=0.007). CONCLUSIONS: Our study has shown that 17ß-estradiol improved myocardial diastolic function, prevented myocardial energy dysregulation, and reduced myocardial oxidative stress in cTnT-Q92 mice.

Shexiang Tongxin dropping pill attenuates atherosclerotic lesions in ApoE deficient mouse model.

ETHNOPHARMACOLOGICAL RELEVANCE: Shexiang Tongxin dropping pill (STDP) is a formulation of Traditional Chinese Medicine mainly used for clinical treatment of stable angina pectoris in China. AIM OF THE STUDY: To investigate the effects and mechanisms of STDP treatment on atherosclerosis. MATERIALS AND METHODS: ApoE deficient (ApoE(-/-)) mice were utilized to evaluate the effect of STDP treatment (30 mg/kg/day) on atherosclerotic lesions. Histopathological features of atherosclerotic lesions, serum levels of lipid proteins, parameters of oxidative stress and pro-inflammatory cytokines were measured by H&E staining, Masson's trichrome staining and ELISA, respectively. Real-time PCR analyses were performed to examine the aortic expression of atherosclerosis-associated microRNAs. RESULTS: The STDP treatment resulted in attenuated atherosclerotic lesion manifested by reduced lipid deposition, fibrosis and oxidative stress. It also led to increase in serum levels of GSH and SOD, decrease in MDA, decrease in CHO, TG, LDL, ox-LDL and increase in HDL, respectively. Additionally, the levels of pro-inflammatory cytokines including IL-2, IL-6, TNF-α and γ-IFN were markedly reduced by STDP treatment. Furthermore, STDP treatment was associated with a significant reduction in the aortic expression of miR-21a, miR-132, miR-126a, miR-155 and increased expression of miR-20a. CONCLUSION: Our results demonstrated for the first time that STDP attenuated atherosclerotic lesions in ApoE(-/-) mouse model. Moreover, STDP treatment exhibited multi-targeting effects on pathological, biochemical and molecular aspects of atherosclerosis implicating lipid regulation, fibrosis, inflammation and oxidative stress. Findings from the current study warrant further evaluation of the clinical application of STDP in atherosclerosis treatment.

Preliminary study of quercetin affecting the hypothalamic-pituitary-gonadal axis on rat endometriosis model.

In this study, the endometriosis rats model was randomly divided into 6 groups: model control group, ovariectomized group, Gestrinone group, and quercetin high/medium/low dose group. Rats were killed after 3 weeks of administration. The expression levels of serum FSH and LH were detected by ELISA. The localizations and quantities of ERα, ERß, and PR were detected by immunohistochemistry and western blot. The results showed that the mechanism of quercetin inhibiting the growth of ectopic endometrium on rat endometriosis model may be through the decreasing of serum FSH and LH levels and then reducing local estrogen content to make the ectopic endometrium atrophy. Quercetin can decrease the expression of ERα, ERß, and PR in hypothalamus, pituitary, and endometrium, thereby inhibiting estrogen and progesterone binding to their receptors to play the role of antiestrogen and progesterone.

Bidirectional regulation of angiogenesis and miR-18a expression by PNS in the mouse model of tumor complicated by myocardial ischemia.

BACKGROUND: Panax Notoginseng Saponins (PNS) is the major class of active constituents of notoginseng, a natural product extensively used as a therapeutic agent in China. Tumor when accompanied by cardiovascular disorders poses a greater challenge for clinical management given the paradoxical involvement of angiogenesis, therefore gaining increased research attention. This study aim to investigate effects of PNS and its activity components in the mouse model of tumor complicated with myocardial ischemia. METHODS: Tumor complexed with myocardial ischemia mouse model was first established, which was followed by histological and immunohistochemistry examination to assess the effect of indicated treatments on tumor, myocardial ischemia and tissue specific angiogenesis. MicroRNA (miRNA) profiling was further carried out to identify potential miRNA regulators that might mechanistically underline the therapeutic effects of PNS in this complex model. RESULTS: PNS and its major activity components Rg1, Rb1 and R1 suppressed tumor growth and simultaneously attenuated myocardial ischemia. PNS treatment led to decreased expression of CD34 and vWF in tumor and increased expression of these vascular markers in heart. PNS treatment resulted in reduced expression of miR-18a in tumor and upregulated expression of miR-18a in heart. CONCLUSIONS: Our data demonstrated for the first time that PNS exerts tissue specific regulatory effects on angiogenesis in part through modulating the expression of miR-18a, which could be responsible for its bidirectional effect on complex disease conditions where paradoxical angiogenesis is implicated. Therefore, our study provides experimental evidence warranting evaluation of PNS and related bioactive component as a rational therapy for complex disease conditions including co-manifestation of cancer and ischemic cardiovascular disease.

Panax notoginseng saponins (PNS) inhibits breast cancer metastasis.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax notoginseng (Burkill) F.H. Chen (Araliaceae) has been extensively used as a therapeutic agent to treat a variety of diseases. Panax notoginseng saponins (PNS) consist of major therapeutically active components of Panax notoginseng. PNS inhibit the growth of a variety of tumor cells in vitro and in vivo. The aim of the study is to investigate the effects and underlying mechanisms of PNS on breast cancer metastasis. MATERIALS AND METHODS: 4T1 cell, a highly metastatic mouse breast carcinoma cell line, was utilized for in vitro and in vivo assays. In vitro assays were first performed to examine the effects of PNS on 4T1 cell viability, migration and invasion, respectively. Real-time PCR analyses were also performed to examine the effects of PNS on the expression of genes associated with tumor metastasis. The effect of PNS on 4T1 tumor cell metastasis was further assessed in spontaneous and experimental metastasis models in vivo. RESULTS: PNS treatment exhibited a dose-dependent effect on impairing 4T1 cell viability in vitro. However, when examined at a lower dose that did not affect cell viability, the migration and invasion of 4T1 cell was remarkably inhibited in vitro. Meanwhile, PNS treatment led to upregulated expression of genes known to inhibit metastasis and downregulated expression of genes promoting metastasis in cultured 4T1 cells. These results suggested a selective effect of PNS on 4T1 migration and invasion. This hypothesis was further addressed in 4T1 metastasis models in vivo. The results showed that the lung metastasis was significantly inhibited by PNS treatment in both spontaneous and experimental metastasis models. CONCLUSION: Taken together, our results demonstrated an inhibitory effect of PNS on 4T1 tumor metastasis, warranting further evaluation of PNS as a therapeutic agent for treating breast cancer metastasis.