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Esophageal cancer (EC) is the sixth leading cause of cancer-related mortality worldwide, with the incidence gradually increasing each year. Therefore, further clarifying the mechanism underlying the development of EC may be beneficial for identifying novel biomarkers and targets for its treatment. The present study aimed to determine the functional roles of glioblastoma-amplified sequence (GBAS), a newly identified gene that has been reported to play crucial roles in multiple types of cancer, including in the malignant behavior of EC cells, such as cell viability, colony formation, cell apoptosis and cell cycle progression. The results of the present study revealed that, in vitro, the knockdown of GBAS significantly suppressed cell viability and colony formation in TE-1 and KYSE-150 cell lines, using a Celigo cell count analysis and colony formation assay respectively, whereas the apoptotic rate of EC cells was significantly increased by the knockdown of GBAS using Annexin V APC staining. Furthermore, following GBAS knockdown, the cell cycle progression of TE-1 and KYSE-150 cells was arrested in the G1 phase using PI staining. In conclusion, the findings of the present study suggested that GBAS may serve a role in EC by regulating cell viability, apoptosis and cell cycle progression.

Disruption of the intestinal mucosal barrier integrity is a pathogenic process in inflammatory bowel disease (IBD) development, and is therefore considered a drug discovery target for IBD. The well-known traditional Chinese formulation Qing Hua Chang Yin (QHCY) has been suggested as a potential therapeutic agent for the treatment of ulcerative colitis. However, the possible underlying molecular mechanisms regarding its therapeutic effect remain unclear. Consequently, the present study investigated the effects of QHCY on lipopolysaccharide (LPS)-induced loss of intestinal epithelial barrier integrity in vitro using the Caco-2 cell model of intestinal epithelium. QHCY reversed the LPS-induced decrease in transepithelial electrical resistance and significantly alleviated the increased fluorescently-labeled dextran 4 flux caused by LPS. Moreover, QHCY upregulated the mRNA and protein expression levels of occludin, zona occludens-1 and claudin-1 in LPS-exposed Caco-2 cells. In conclusion, QHCY was able to protect intestinal epithelial barrier integrity following an inflammatory insult; the protective effects of QHCY may be mediated by modulation of the expression of tight junction proteins.

Curcumin (Cur) has been reported to function as an antioxidant and anti-inflammatory agent and to play a role in anti-atherosclerosis. The present study aimed to explore the protective effect of Cur on hypoxia/reoxygenation (H/R) injury. The morphological changes in H9c2 cardiomyocytes were observed under an inverted microscope. Cell viability was determined by Cell Counting Kit-8 (CCK-8). Lactate dehydrogenase (LDH) level, malondialdehyde (MDA) level and the antioxidant superoxide dismutase (SOD) activity were determined by corresponding kits. Apoptosis and reactive oxygen species (ROS) levels were determined by flow cytometry. Endoplasmic reticulum (ER) stress-related factors, which were examined by quantitative real-time polymerase chain reaction (qPCR) and western blot analysis, included 78-kDa glucose-regulated protein (GRP78) and C/EBP homologous protein (CHOP). Extracellular signal regulating kinase 1/2 (ERK1/2), p38, c-Jun NH2-terminal kinase (JNK) and the phosphorylation levels of key proteins in the mitogen-activated protein kinase (MAPK) signaling pathway were all determined by western blot analysis. Compared to the control group, the cell morphology of the H9c2 cells was obviously altered upon H/R. Cell viability was significantly decreased, while apoptosis was significantly increased by H/R. We also observed that the levels of LDH and MDA were elevated and the activity of SOD was decreased in the H/R group. Notably, LDH, MDA and SOD levels were reversed following treatment with Cur; while apoptosis and ROS levels in the H/R injury group were decreased by Cur. H/R injury-triggered ER stress and the MAPK signaling pathway were suppressed by Cur. These results demonstrated that Cur has a protective effect on cardiomyocytes via suppression of ER stress and the MAPK pathway.

Mesenchymal stem cell‑conditioned medium (MSC‑CM) contains various cytokines (osteopontin and macrophage colony stimulating factor 1) secreted by MSCs and may modulate the immune response in tubulointerstitial fibrosis. The aim of the present study was to investigate whether MSC‑CM treatment may affect B cell‑dependent immune responses, which have previously been reported to facilitate the renal fibrotic processes following unilateral ureteral obstruction (UUO). In the present study, histological analysis, flow cytometry, western blotting and reverse transcription‑quantitative polymerase chain reaction were performed. MSC‑CM treatment was observed to impede renal infiltration of B lymphocytes and the expression of CC chemokine ligand‑2. Additionally, UUO suppressed the subsequent recruitment of monocytes/macrophages to the kidney, limited local inflammation and attenuated renal fibrosis. The findings of the present study identified a potential mechanism of MSC‑CM in ameliorating the UUO‑kidney.

Combination chemotherapy with chemosensitizers can exert synergistic therapeutic effects, reduce toxicity, and delay the induction of drug resistance. In the present study, the antitumor effects were investigated, and the possible underlying mechanisms of kaempferol combined with 5‑fluorouracil (5‑FU) in colorectal cancer cells were explored. HCT‑8 or HCT‑116 cells were treated with various concentrations of kaempferol and/or 5‑FU for the indicated time‑points. An MTT assay was used to determine cell viability, whereas the synergistic effects were assessed by calculating the combination indices of kaempferol and 5‑FU. Annexin V analysis and Hoechst staining were used to determine cell apoptosis. q‑PCR and western blotting were performed to determine the expression levels of Bax, Bcl‑2, thymidylate synthase (TS), PTEN, PI3K, AKT, and p‑AKT. The combination of kaempferol and 5‑FU was determined to be more effective in inhibiting cell viability than either of the agents alone. The inhibition of tumors in response to kaempferol and 5‑FU was associated with the reduction in proliferation ability and stimulation of apoptosis. The protein results indicated that kaempferol and 5‑FU could significantly upregulate the expression levels of Bax and downregulate the expression levels of Bcl‑2 and TS. Furthermore, the combination treatment greatly inhibited the activation of the PI3K/Akt pathway, suggesting the involvement of this pathway in the synergistic effects. The present study demonstrated that kaempferol has a synergistic effect with 5‑FU by inhibiting cell proliferation and inducing apoptosis in colorectal cancer cells via suppression of TS or attenuation of p‑Akt activation. The combination of kaempferol and 5‑FU may be used as an effective therapeutic strategy for colorectal cancer.

Ursolic acid, a pentacyclic triterpene compound with low toxicity and easy availability, has a variety of biological activities, including antitumor, antioxidant, antihepatitis, anti‑inflammatory and antibacterial effects. The present study aimed to investigate the renoprotective effects of ursolic acid on ischemia/reperfusion‑induced acute kidney injury (I/R‑IAKI) in rats associated with its antioxidant and anti‑inflammatory effects, as well as interference with the signal transducer and activator of transcription (STAT)3/nuclear factor (NF)‑κB signaling pathway. The present study demonstrated that pre‑treatment with ursolic acid significantly increased renal functioning and attenuated increases of serum angiotensin II levels in rats subjected to I/R‑IAKI. In addition, I/R‑IAKI‑induced inflammation and oxidative stress were significantly reduced by pre‑treatment with ursolic acid. Furthermore, ursolic acid significantly suppressed the upregulation of STAT3, NF‑κB and caspase‑3 activities in rats following I/R‑IAKI. These results indicated that ursolic acid may be a potential drug for reducing I/R‑IAKI through suppression of inflammation and oxidative stress damage, as well as modulation of STAT3 and NF‑κB activities.

Angiogenesis is an essential process of cancer progression and is regulated by multiple intracellular signaling pathways, including signal transducer and activator of transcription 3 (STAT3) and sonic hedgehog (SHH). Thus, these pathways have become a promising target for anti‑cancer therapeutic strategies. Oleanolic acid (OA) is an active compound present in various herbal medicines, which have been used historically for the clinical treatment of various types of human malignancies, including colorectal cancer (CRC). The present study used a CRC mouse xenograft model and human umbilical vein endothelial cells (HUVECs) to evaluate the effect of OA on tumor angiogenesis and on the activation of the STAT3 and SHH signaling pathways. It was determined that OA treatment significantly inhibited tumor growth and reduced intratumoral microvessel density (MVD) in CRC mice. In addition, OA treatment inhibited the proliferation, migration and tube formation in HUVECs, in a dose and time-dependent manner. Furthermore, OA markedly suppressed the activation of the STAT3 and SHH signaling pathways and inhibited the expression of the pro‑angiogenic vascular endothelial growth factor A and basic fibroblast growth factor, two important target genes of the aforementioned signaling pathways. Therefore it is suggested that inhibition of tumor angiogenesis via the suppression of multiple signaling pathways may be one of the underlying mechanisms by which OA exerts its anti-cancer effect.

The aim of the present study was to investigate whether the cyclic adenosine 3',5'‑monophosphate (cAMP)/protein kinase A(PKA)/cAMP‑responsive element binding protein (CREB) signal transduction pathway triggered by γ‑aminobutyric acid class B (GABA(B)) receptor activation is involved in neuroprotection against ischemia and behavioral recovery induced by opposing needling (ON). A total of 80 rats were randomly divided into four groups: A sham operation group, an ischemia group, an ON group and an ON group effectively inhibited by the GABA(B) receptor antagonist, CGP35384 (n=20/group). The behavior of the rats was assessed by their neurological deficit score, whereas the impairment of gait was examined using the CatWalk system. The volume of cerebral infarction was examined upon treatment with 2,3,5‑triphenyltetrazolium chloride. The expression levels of CREB, GABA(B1) and GABA(B2) were examined by western blotting and reverse transcription‑quantitative polymerase chain reaction, and the activity of adenylyl cyclase (AC), cAMP and PKA in the serum was detected using an enzyme‑linked immunosorbent assay. In the present study, in comparison with other groups, the ON group exhibited a reduced score for the neurological deficit, the stride length and swing speed were improved, and the volume of infarction was reduced. However, these effects were reversed upon administration of CGP35384. Additionally, the expression levels of CREB, GABA(B1) and GABA(B2) were increased in the ON group. The levels of AC, cAMP and PKA in the serum were also increased in the ON group, whereas the addition of CGP35384 reversed these effects. The results of the present study demonstrated that ON markedly protected the brain against transient cerebral ischemic injury, and this effect was possibly mediated by the activation of the GABAB/cAMP/PKA/CREB signal transduction pathway. These findings implied that ON may be a potential therapeutic method for treating stroke.

Interleukin‑6 (IL‑6)/signal transducer and activator of transcription 3 (STAT3) pathway plays essential roles in the development of inflammatory diseases including ulcerative colitis (UC). Therefore, suppression of IL‑6/STAT3 signaling provides a promising therapeutic strategy in UC. Pien Tze Huang (PZH), a well‑known traditional Chinese formula, has been used in China and Southeast Asia for centuries as a folk remedy for various inflammatory diseases. However, the molecular mechanisms of its anti‑inflammatory effects remain to be elucidated. In the present study, we generated a mouse colitis model by using dextran sulfate sodium (DSS) and evaluated the therapeutic efficacy of PZH against UC by observing the clinical manifestations. We found that PZH obviously alleviated DSS‑induced colitis symptoms, including body weight loss, rectal bleeding and stool consistency. In addition, administration of PZH profoundly prevented DSS‑induced colon shortening, and ameliorated colonic histopathological changes such as mucosal ulceration, infiltration of inflammatory cells, crypt distortion and hyperplastic epithelium. Moreover, PZH markedly inhibited the serum level of the inflammatory biomarker serum amylase A (SAA) in UC mice. Furthermore, PZH treatment significantly inhibited DSS‑induced expression of IL‑6 in colon tissues. Finally, the increased phosphorylation level of STAT3, induced either by DSS in experimental mice or by IL‑6 in the differentiated human colorectal carcinoma cells, was significantly suppressed by PZH. These results suggest that the inhibition of IL‑6/STAT3 signaling is a potential mechanism by which PZH is used in the treatment of UC.

Hedyotis diffusa Willd (HDW) is a major component frequently used in Traditional Chinese Medicine for the clinical treatment of colorectal cancer (CRC) and its associated drug resistance. However, the underlying mechanism of HDW circumventing drug resistance of cancer cells remains to be elucidated. Cancer cell resistance to apoptosis and activation of the phosphatidylinositol‑3‑kinase (PI3K)/protein kinase B (AKT) signaling pathway have been implicated as major factors in the acquired resistance to chemotherapeutic anti‑cancer drugs. The present study investigated the effect and mechanisms of action of ethanol extract of Hedyotis diffusa Willd (EEHDW) on the proliferation and apoptosis of CRC 5‑fluorouracil (5‑FU) resistant (HCT‑8/5‑FU) cells. CRC HCT‑8/5‑FU cell viability following treatment with EEHDW was determined using MTT and colony formation assay. In addition, Annexin V/propidium iodide staining with flow cytometry analysis and 4',6‑diamidino‑2‑phenylindole staining were used to determine the apoptosis of drug‑resistant cancer cells following treatment with EEHDW. The mRNA and protein expression levels of B cell leukemia/lymphoma (Bcl‑2), Bcl‑2 associated X (Bax), cyclin dependent kinase 4 (CDK4), cyclin D1 and p21 were evaluated using reverse transcription‑polymerase chain reaction and western blot analysis, respectively. Furthermore, activation of the PI3K/AKT signaling pathway and expression of phosphatase and tensin homolog (PTEN), PI3K, AKT and phosphorylated (p)‑AKT were determined. EEHDW significantly reduced cell viability, inhibited cell colony formation and promoted apoptosis of HCT‑8/5‑FU cells. Furthermore, EEHDW significantly downregulated the expression of Bcl‑2, cyclin D1 and CDK4 and upregulated the expression of Bax and p21. In addition, EEHDW inhibited the activation of the PI3K/AKT pathway by increasing expression of PTEN and suppressing the expression of PI3K and p‑AKT. The present study provided the first direct evidence that EEHDW may overcome drug‑resistance in human CRC cells by inhibiting PI3K/AKT signaling pathway and provides a basis for the improved therapeutic use of HDW in the clinical treatment of cancer.

The widely-used Chinese medicinal herb Hypericum japonicum, also known as Hypericum japonicum Thunb or Tianjihuang, displays potent anti‑carcinogenic effects against liver cancer. However, the molecular mechanism underlying the therapeutic effects of Hypericum japonicum remains to be elucidated. The present study investigated the in vivo efficacy of ethyl acetate extract of Hypericum japonicum (EAEHJ) against tumor growth in an H22 cell‑bearing liver cancer mouse model. Treatment with EAEHJ significantly reduced tumor weight, but had no effect on murine body weight. The results of the present study also showed that EAEHJ induced H22 cell apoptosis in vivo. In addition, the anti‑carcinogenic effects of EAEHJ were investigated in vitro. The results of the present study demonstrate that both phospholipid asymmetry in the plasma membrane and mitochondrial membrane potential were deregulated in HepG2 human hepatoma cells, following treatment with EAEHJ. Treatment with EAEHJ also increased the ratio of pro‑apoptotic B‑cell lymphoma 2 (Bcl‑2)‑associated X protein (Bax) to anti‑apoptotic Bcl‑2, and activated the caspase‑9 signaling pathway. These results suggest that EAEHJ is able to trigger the apoptosis of liver cancer cells via the mitochondria-dependent pathway.

The signal transducer and activator of transcription 3 (STAT3) pathway is one of the main growth factor‑mediated signal transduction pathways and is closely associated with the occurrence and development of benign prostatic hyperplasia (BPH). Qianliening capsules (QC) have significant therapeutic effects on BPH; however, the precise mechanism underlying its anti‑BPH activity remains to be elucidated. To further elucidate the molecular mechanism of the therapeutic effect of QC on BPH, the present study used epidermal growth factor (EGF), which has a role in the pathogenesis of BPH, to stimulate the growth of human prostate WPMY‑1 cells and activate the STAT3 pathway in the WPMY‑1 cells. The cell viability was determined using an MTT assay and the cell morphology was observed by phase‑contrast microscopy. Fluorescence activated cell sorting analysis with Annexin‑V/propidium iodide (PI) staining and PI staining were performed to examine cell apoptosis and the cell cycle. The activation of caspase‑9 and ‑3 were evaluated by colorimetric assay. STAT3 phosphorylation and transcriptional activity were detected by western blot analysis and the luciferase gene reporter, respectively. The mRNA and protein expression levels of B‑cell lymhoma 2 (Bcl‑2), Bcl‑2‑associated X protein (Bax), cyclin D1, cyclin‑dependent kinase 4 (CDK4) and p21 were measured by reverse transcription quantitative polymerase chain reaction and western blot analysis, respectively. In the present study, QC was found to significantly and dose‑dependently inhibit the EGF‑stimulated growth of WPMY‑1 cells, as evidenced by QC‑induced cell -morphological changes and a reduction in cell viability. In addition, QC treatment markedly induced the activation of caspase‑9 and ‑3. QC treatment also inhibited the EGF‑mediated increase of STAT3 phosphorylation levels and transcriptional activity in WPMY‑19 cells, accompanied by downregulation of the expression of Bcl‑2, cyclin D1 and CDK4 and upregulation of the expression of Bax and p21. These results suggested that QC effectively inhibited the proliferation and promoted the apoptosis of human prostate cells via modulation of the STAT3 signaling pathway and its target genes, which is likely to be one of the mechanisms underlying its activity in BPH treatment.

The plant Rubus alceaefolius Poir is used as a hepatic protectant in Traditional Chinese Medicine. The aim of the present study was to confirm the protective effect of the total alkaloids of Rubus alceaefolius Poir (TARAP) on the liver and to evaluate the potential molecular mechanisms associated with adipocytokines underlying non-alcoholic fatty liver disease (NAFLD) in rats. To generate the NAFLD model, Sprague-Dawley rats were administered a high-fat diet and following 12 weeks of model construction, rats were orally treated with a positive control drug and different doses of TARAP daily for 28 days. The rats were then sacrificed and the livers were collected to evaluate the liver index (LI) and observe histological changes by hematoxylin and eosin (H&E) staining. The secretion levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum were examined by ELISA. Finally, the expression levels of leptin (LEP), resistin and adiponectin (APN) in liver tissues were determined by immunohistochemistry (IHC). The results demonstrated that, in the group treated with methionine and choline bitartrate tablets and in the groups treated with different doses of TARAP, there was a significant reduction in the LI (P

Nogo-A has been identified as an inhibitor of neurite outgrowth specific to the central nervous system. However, little is known about the role of Nogo-A in hepatocellular carcinoma (HCC), the most common primary malignant tumor with a high mortality rate. This study aimed to investigate the role of endogenous Nogo-A in human liver cancer cells. Reverse transcription polymerase chain reaction was used to detect the expression of Nogo-A in four liver cancer cell lines. A lentivirus vector was then constructed to mediate RNA interference (RNAi) targeting of Nogo‑A (LV‑Nogo-A‑siRNA) and was confirmed to successfully suppress the expression of the Nogo-A gene in SMMC-7721 cells. Furthermore, Nogo-A was observed to be highly expressed in liver cancer cell lines. RNAi of Nogo-A using the LV‑Nogo-A‑siRNA construct significantly decreased Nogo-A protein expression and specifically inhibited the growth of SMMC-7721 cells. This growth inhibitory effect may be attributed to an increase in G2/M phase arrest and apoptosis in SMMC-7721 cells containing Nogo-A‑siRNA. The results of this study demonstrate that Nogo-A may represent a novel therapeutic target for the treatment of liver cancer, in addition to its potent roles in neural systems.

Colorectal cancer (CRC) is one of the most common malignant tumors of the digestive tract, and threatens the survival and health of patients with CRC. Chemotherapy remains one of the main therapeutic approaches for patients with CRC; however, drug resistance limits the long‑term use. CRC cells with multi‑drug resistance (MDR) exhibit increased survival times and metastatic potential, which may lead to the recurrence and metastasis of CRC. In addition, MDR is one of the major causes of chemotherapy failure in clinical treatment. Hedyotis diffusa Willd (HDW) has been used in the treatment of inflammation‑associated diseases and malignant tumors, including CRC. The authors previously demonstrated that HDW could reverse MDR in CRC cells; however, its underlying mechanism, particularly in MDR‑associated metastasis, remains to be elucidated. In the present study, the drug‑resistant CRC cell line HCT‑8/5‑fluorouracil (5‑FU) was used to investigate the effect of HDW on the growth and metastasis of cancer cells. Cell viability was assessed using the MTT assay. Cell adhesion potential was evaluated using adhesion experiments. Cell migration was assessed using wound healing and Transwell assays. The mRNA and protein expression levels of crucial factors in the transforming growth factor‑β (TGF‑β) signaling pathway, including TGF‑β, Mothers against decapentaplegic homolog 4 (SMAD4), neural (N)‑cadherin, and epithelial (E)‑cadherin, were analyzed using the reverse transcription‑semi‑quantitative polymerase chain reaction and western blotting, respectively. The results demonstrated that the HCT‑8/5‑FU cell line was more resistant to 5‑FU and thus can be used as the resistant cell model. HDW was able to inhibit the viability, and adhesive, migratory and invasion potential of the HCT‑8/5‑FU cells. In addition, HDW was able to downregulate the expression of TGF‑β, SMAD4 and N‑cadherin, and upregulate E‑cadherin, at the gene and protein level. In conclusion, the results demonstrated that HDW may suppress the metastasis of 5‑FU‑resistant CRC cells via regulation of the TGF‑β signaling pathway, which was also considered to be one of the underlying mechanisms of its anti‑CRC effect.

The aim of this study was to evaluate the therapeutic efficacy of Qianliening capsule (QC) against benign prostatic hyperplasia (BPH) in vivo in a BPH rat model, as well as to investigate the effects of QC on prostatic cell apoptosis and the possible molecular mechanisms mediating its anti-BPH activity. Fifty male Sprague‑Dawley (SD) rats were randomly classified into five groups. The rats of the four groups were castrated and subcutaneously injected with testosterone propionate to generate BPH. One week after model establishment, BPH rats were orally administrated with various doses of QC daily for 28 days. The prostatic tissues from BPH rats were collected to evaluate prostatic index (PI). The histological changes of prostate were observed by hematoxylin and eosin staining. TUNEL analysis was performed to examine cell apoptosis. The mRNA expression of Bcl-2 and Bax in prostatic tissues was determined by reverse transcription‑polymerase chain reaction (RT-PCR). The protein expression of Bcl-2, Bax and cleaved caspase 3 were examined by immunohistochemistry. Administration with QC significantly decreased PI in a dose-dependent manner (P

Shexiang Tongxin Dropping Pill (STP) is an established traditional Chinese medicine that is widely used for the treatment of ischemic heart disease (IHD), although its mechanisms remain unclear. The present study investigated the protective effects of STP following pituitrin (PTT)‑induced myocardial ischemia in rats. ST‑segment elevation, blood rheology, and the serum levels of creatine kinase‑MB (CK‑MB) and lactate dehydrogenase (LDH) were measured. Following heart excision, histological analysis using hematoxylin and eosin and terminal deoxynucleotidyl transferase dUTP nick end labeling were performed. The mRNA expression levels of B‑cell lymphoma 2 (Bcl‑2) and Bcl‑2‑associated X protein (Bax) were determined using reverse transcription‑quantitative polymerase chain reaction, and their protein expression was detected using immunohistochemistry. The results demonstrated that STP treatment protected against ST elevation, lowered whole blood viscosity, and reduced the serum levels of CK‑MB and LDH following acute myocardial ischemia. In addition, STP treatment restored the histopathological change following PTT‑induced myocardial ischemia, and resulted in downregulated expression of Bax and upregulated expression of Bcl‑2 in myocardial tissue. The present study demonstrates the cardioprotective ability of STP in a rat model of myocardial ischemic injury, which may be attributed to its anti‑apoptotic properties. The cardioprotective properties of STP require further investigation to determine whether it may be used for the clinical treatment of IHDs.

Non‑alcoholic fatty liver disease (NAFLD) is a widespread health concern, and there is currently insufficient understanding regarding its pathogenesis and treatment. The present study aimed to explore the effects of Artemisia capillaris formula (ACF) on high‑fat diet‑induced hepatic steatosis and fatty acid‑induced intracellular lipid accumulation, by micro (mi)RNA regulation. A total of 72 Sprague‑Dawley rats were divided into six groups (n=12/group). One group was designated as the control group and fed a normal diet, and the remaining five groups were allowed ad libitum access to a high‑fat diet for eight weeks, in order to establish an NAFLD rat model. The rats were subsequently administered polyene phosphatidylcholine (PP; 0.076 g/kg body weight/day), low dose of ACF (0.462 g/kg body weight/day), middle dose of ACF (0.924 g/kg body weight/day) or high dose of ACF (1.848 g/kg body weight/day) intragastrically for four weeks. HepG2 human hepatocellular carcinoma cells were treated with oleic acid and palm acid, followed by treatment with various concentrations of ACF. Serum alanine transaminase (ALT), aspartate aminotransferase (AST), triglycerides (TG), total cholesterol (TC), high‑density lipoprotein cholesterol (HDL‑C), low‑density lipoprotein cholesterol (LDL‑C), and steatotic HepG2 human liver carcinoma cell TC and TG levels were measured. ACF and PP treatments attenuated high‑fat diet‑induced hepatic steatosis and fatty acid‑induced intracellular lipid accumulation. A modified high‑fat diet significantly increased ALT, AST, TG, TC, LDL‑C levels and decreased HDL‑C levels. Treatment with ACF and PP abrogated the increase in liver enzymes and TG, TC and LDL‑C levels, but did not influence HDL‑C levels in a high‑fat diet induced rat model of steotosis. Steatotic HepG2 cells exhibited significantly increased levels of both TG and TC. Treatment with ACF significantly decreased TC and TG levels in vivo, and ACF and PP treatment decreased the expression levels of fatty acid synthase (FASN) and increased miR‑122 in vivo and in vitro. In conclusion, these results suggested that ACF may inhibit hepatic steatosis via miR‑122‑induced downregulation of FASN in vivo and in vitro.

Angiogenesis, which has a critical role in human tumor growth and development, is tightly regulated by the Notch signaling pathway. Total alkaloids are active components of the plant Rubus alceifolius Poir, which is used for the treatment of various types of cancer. A previous study by our group showed that the total alkaloids of Rubus alceifolius Poir (TARAP) induced hepatocellular carcinoma (HCC) cell apoptosis through the activation of the mitochondria-dependent pathway in vitro and in vivo, as well as inhibited angiogenesis in a chick embryo chorioallantoic membrane model. In the present study, to further analyze the specific mechanisms underlying the antitumor activity of TARAP, a HCC xenograft mouse model was used to assess the effect of TARAP on angiogenesis in vivo. TARAP was found to suppress the expression of vascular endothelial growth factor (VEGF) A and VEGF receptor-2 in tumor tissues, which resulted in the inhibition of tumor angiogenesis. In addition, TARAP treatment was observed to inhibit the expression of Notch1, delta-like ligand 4 and jagged 1, which are key mediators of the Notch signaling pathway. The present study identified that the inhibition of tumor angiogenesis through the suppression of the Notch signaling pathway may be one of the mechanisms through which TARAP may be effective in the treatment of cancer.

The transforming growth factor-β (TGF-β) signaling pathway exhibits an important role in cancer invasion and metastasis. Excessive expression of TGF-β activates Smad4, leading to the upregulation of downstream metastasis-associated genes. Thus, the inhibition of the TGF-β/Smad4 signaling pathway may be a novel strategy for treatment of cancer metastasis. Baicalein, a flavonoid derived from the root of Scutellaria baicalensis, has been reported to exert strong anti-tumor activity towards various types of cancer. In the present study the effect of baicalein on migration and invasion of cancer cells was evaluated using wound-healing and Transwell assays. In order to investigate the possible molecular mechanisms of the anti-metastatic effects of baicalein, quantitative polymerase chain reaction (qPCR) and western blot analyses were performed to examine the effect on the expression of TGF‑β, Smad4, N-cadherin, vimentin, ZEB1 and ZEB2. It was determined that baicalein inhibited the migration and invasion of AGS cells by suppressing the TGF-β/Smad4 signaling pathway. In addition, baicalein treatment reduced the expression of the metastasis-associated N-cadherin, vimentin, ZEB1 and ZEB2, downstream target genes of the TGF‑β/Smad4 signaling pathway. Collectively, these results suggest that inhibition of the metastasis of cancer cells via inactivation of TGF-β/Smad4 signaling is one of the mechanisms by which baicalein may treat cancer.

Accumulating evidence suggests that a small population of cells termed cancer stem cells (CSCs) are crucial in tumor development and drug resistance, leading to cancer relapse and metastasis and eventually the failure of clinical cancer treatment. Therefore, targeting CSCs is a promising approach for anticancer therapies. Due to the drug resistance and adverse effects of currently used chemo- therapies, traditional Chinese medicines (TCM) have recently received attention due to the relatively few side-effects. Thus, they have been used as important alternative remedies for various diseases, including cancer. Pien Tze Huang (PZH), a well-known TCM formula that was first prescribed more than 450 years ago in the Ming Dynasty, has been used in China and Southeast Asia for centuries as a folk remedy for various types of cancer. Previously, it was reported that PZH inhibits colon cancer growth via the promotion of cancer cell apoptosis and inhibition of cell proliferation and tumor angiogenesis, which is probably mediated by its regula- tory effect on multiple intracellular signaling pathways. To elucidate the mechanism of the tumoricidal activity of PZH, the aim of the present study was to investigate the effect of PZH on CSCs that were isolated as the side population (SP) from the HT-29 colorectal cancer cell line. The results demonstrated that PZH significantly and dose-dependently reduced the percentage of the colorectal cancer stem-like SP cells, decreased the viability and sphere-forming capacity of HT-29 SP cells, indicating that PZH is potent in suppressing the growth of colorectal cancer stem cells. Moreover, PZH treatment in HT-29 SP cells markedly inhibited the mRNA levels of ABCB1 and ABCG2, which are members of the ABC transporter superfamily, thereby contributing to the SP phenotype and multi-drug resistance. Findings of the present study suggest that inhibiting the growth of CSCs is a potential mechanism by which PZH can be used in cancer treatment.

Scutellaria barbata D. Don (SB) has long been used as a major component in numerous Chinese medical formulas to clinically treat various types of cancer. Previously, we reported that the extracts of SB were able to suppress colon cancer growth in vivo and in vitro, possibly by inducing cancer cell apoptosis and inhibiting cell proliferation and tumor angiogenesis. However, the anticancer mechanisms of its bioactive ingredients remain largely unclear. In the present study, using three human colon cancer cell lines SW620, HT-29 and HCT-8, the antitumor effect of different solvent fractions of SB were evaluated and the potential underlying molecular mechanisms were investigated. Using an 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, it was revealed that the chloroform fraction of SB (ECSB) exhibited the most potent inhibitory effect on the growth of all three colon cancer cell lines and SW620 cells exhibited the most sensitive response to ECSB treatment (IC50=65 µg/ml). In addition, by performing fluorescence-activated cell sorting, transmission electron microscopy and colony formation assays, it was observed that ECSB significantly induced apoptosis and inhibited proliferation in SW620 cells in a dose-dependent manner. Furthermore, ECSB treatment resulted in the upregulation of the pro-apoptotic Bax/Bcl-2 ratio and a decrease in the expression of the pro-proliferative cyclin D1 and cyclin-dependent kinase 4. The results from the present study may provide a scientific foundation for the development of novel anticancer agents from the bioactive ingredients in the ECSB.

Cognitive impairment is a serious mental deficit following stroke that severely affects the quality of life of stroke survivors. Nuclear factor‑κB (NF-κB)-mediated neuronal cell apoptosis is involved in the development of post-stroke cognitive impairment; therefore, it has become a promising target for the treatment of impaired cognition. Acupuncture at the Baihui (DU20) and Shenting (DU24) acupoints is commonly used in China to clinically treat post‑stroke cognitive impairment; however, the precise mechanism of its action is largely unknown. In the present study, we evaluated the therapeutic efficacy of electroacupuncture against post-stroke cognitive impairment and investigated the underlying molecular mechanisms using a rat model of focal cerebral ischemia-reperfusion (I/R) injury. Electroacupuncture at Baihui and Shenting was identified to significantly ameliorate neurological deficits and reduce cerebral infarct volume. Additionally, electroacupuncture improved learning and memory ability in cerebral I/R injured rats, demonstrating its therapeutic efficacy against post-stroke cognitive impairment. Furthermore, electroacupuncture significantly suppressed the I/R-induced activation of NF-κB signaling in ischemic cerebral tissues. The inhibitory effect of electroacupuncture on NF-κB activation led to the inhibition of cerebral cell apoptosis. Finally, electroacupuncture markedly downregulated the expression of pro-apoptotic Bax and Fas, two critical downstream target genes of the NF-κB pathway. Collectively, our findings suggest that inhibition of NF-κB‑mediated neuronal cell apoptosis may be one mechanism via which electroacupuncture at Baihui and Shenting exerts a therapeutic effect on post-stroke cognitive impairment.

Inhibition of tumor angiogenesis has become an attractive target of anticancer chemotherapy. However, drug resistance and cytotoxicity against non-tumor associated endothelial cells limit the long-term use and the therapeutic effectiveness of angiogenesis inhibitors, thus increasing the necessity for the development of multi-target agents with minimal side effects. Traditional Chinese medicine (TCM) formulas, which have relatively fewer side effects and have been used clinically to treat various types of diseases, including cancer, for thousands of years, are considered to be multi-component and multi-target agents exerting their therapeutic function in a more holistic way. Hedyotis Diffusa Willd (EEHDW) has long been used as an important component in several TCM formulas to treat various types of cancer. Although recently we reported that EEHDW promotes cancer cell apoptosis via activation of the mitochondrial-dependent pathway, the precise mechanism of its tumoricidalactivity still remains to be clarified. In the present study, we investigated the angiogenic effects of the ethanol extract of EEHDW. Cell cycle analysis was perfomed using flow cytometry. Cell viability was analyzed using MTT assay. We found that EEHDW inhibited angiogenesis in vivo in chick embryo chorioallantoic membrane (CAM). In addition, we observed that EEHDW dose- and time-dependently inhibited the prolife-ration of human umbilical vein endothelial cells (HUVEC) by blocking the cell cycle G1 to S progression. Moreover, EEHDW inhibited the migration and tube formation of HUVECs. Furthermore, EEHDW treatment down-regulated the mRNA and protein expression levels of VEGF-A in HT-29 human colon carcinoma cells and HUVECs. Our findings suggest that inhibiting tumor angiogenesis is one of the mechanisms by which EEHDW is involved in cancer therapy.

Millimeter wave (MW) is an electromagnetic wave with a wavelength between 1 and 10 mm and a frequency of 30-300 GHz that causes multiple biological effects, both locally and globally. MW has been widely used in clinical medicine. Although our previous work demonstrated that MW is capable of inhibiting sodium nitroprussiate (SNP)-induced apoptosis in chondrocytes, the precise mechanism of the anti-apoptotic activity remains to be elucidated. The purpose of this study was to investigate the effects of MW in SNP-induced apoptotic chondrocytes. Sprague Dawley rat chondrocytes were isolated and cultured, and the cells were counted. Cell viability was evaluated using MTT assay. Cells were then treated with SNP and MW, and flow cytometry was used to detect apoptosis. Our results showed that MW treatment inhibited a SNP-induced mitochondrion-dependent pathway of apoptosis. MW treatment inhibited the loss of plasma membrane asymmetry (externalization of phosphatidylserine), collapse of mitochondrial membrane potential, and activation of caspase-9 and caspase-3. Taken together, the results indicate that MW inhibits the mitochondrion-dependent pathway of apoptosis in chondrocytes and this may, in part, explain its clinical effect in the treatment of osteoarthritis.

Signal transducer and activator of transcription 3 (STAT3) plays an important role in tumor cell survival and proliferation and thus has become a major focus in the development of anti-cancer therapies. Patrinia scabiosaefolia has been used for the treatment of various types of cancer. However, the precise mechanism of the anti-cancer activity of Patrinia scabiosaefolia remains unclear. In this study, we evaluated the effect of the ethanol extract of Patrinia scabiosaefolia (EEPS) on proliferation and apoptosis in human multiple myeloma U266 cells that persistently express phosphorylated STAT3, and investigated the possible molecular mechanisms mediating its biological effects. We found that EEPS inhibited the phosphorylation of STAT3 in U266 cells. Consequently, the inhibitory effect of EEPS on STAT3 activation resulted in the suppression of cell proliferation and the induction of cell apoptosis. Moreover, EEPS treatment inhibited the expression of cyclin D1 (a promoter of cell proliferation) and Bcl-2 (an inhibitor of apoptosis), two important target genes of the STAT3 signaling pathway. Our findings for the first time demonstrate that Patrinia scabiosaefolia inhibits proliferation and promotes the apoptosis of cancer cells via inhibition of the STAT3 pathway, which may in part explain its anti-cancer activity.

Bone morphogenetic protein 2 (BMP2), a member of the transforming growth factor-β (TGF-β) superfamily, plays a key role in the induction of the differentiation of mesenchymal cells into chondrocytes to form cartilage tissue. However, it is not clear whether BMP2 regulates the proliferation of chondrocytes. In the present study, the effect of BMP2 on the proliferation of chondrocytes and its underlying mechanism was investigated. Chondrocytes isolated from the knee of SD rats were cultured and identified using toluidine blue staining. The second generation chondrocytes were collected and stimulated with or without BMP2 for 48 h. Cell viability was analyzed using the MTT assay. mRNA and protein expression levels of β-catenin, GSK-3β, Dvl1 and Cyclin D1 were detected using real-time RT-PCR and Western blotting, respectively. The cell cycle distribution of the chondrocytes was analyzed by flow cytometry. BMP2 stimulation was found to significantly increase cell viability. In addition, following BMP2 treatment, β-catenin, Cyclin D1 and Dvl1 expression was significantly increased, whereas GSK-3β expression was significantly decreased. Moreover, the percentage proportion of chondrocytes in the G0/G1 phase was significantly decreased, whereas that in the S phase was significantly increased. The results indicate that BMP2 promotes chondrocyte proliferation via the Wnt/β-catenin signaling pathway.