Advancements of Macrophages Involvement in Pathological Progression of Colitis-Associated Colorectal Cancer and Associated Pharmacological Interventions.

Intestinal macrophages play crucial roles in both intestinal inflammation and immune homeostasis. They can adopt two distinct phenotypes, primarily determined by environmental cues. These phenotypes encompass the classically activated pro-inflammatory M1 phenotype, as well as the alternatively activated anti-inflammatory M2 phenotype. In regular conditions, intestinal macrophages serve to shield the gut from inflammatory harm. However, when a combination of genetic and environmental elements influences the polarization of these macrophages, it can result in an M1/M2 macrophage activation imbalance, subsequently leading to a loss of control over intestinal inflammation. This shift transforms normal inflammatory responses into pathological damage within the intestines. In patients with ulcerative colitis-associated colorectal cancer (UC-CRC), disorders related to intestinal inflammation are closely correlated with an imbalance in the polarization of intestinal M1/M2 macrophages. Therefore, reinstating the equilibrium in M1/M2 macrophage polarization could potentially serve as an effective approach to the prevention and treatment of UC-CRC. This paper aims to scrutinize the clinical evidence regarding Chinese medicine (CM) in the treatment of UC-CRC, the pivotal role of macrophage polarization in UC-CRC pathogenesis, and the potential mechanisms through which CM regulates macrophage polarization to address UC-CRC. Our objective is to offer fresh perspectives for clinical application, fundamental research, and pharmaceutical advancement in UC-CRC.

Sarcandra glabra (Thunb.) Nakai alleviates DSS-induced ulcerative colitis by promoting restitution, restoring barrier function, and modulating IL-17/Notch1/FoxP3 pathway in intestinal cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Sarcandra glabra is officially named Zhong Jie Feng as a traditional medicine. In the nationality of Yao and Zhuang, it has been used to treat digestive diseases like stomachache and dysentery. Similarly, in Dai nationality, it has been used to treat intestinal diseases like gastric ulcers. However, the effect and mechanism of S. glabra on experimental ulcerative colitis (UC) are known. AIM OF STUDY: The main objective of this study was to investigate the effect and mechanism of S. glabra on experimental UC. MATERIALS AND METHODS: The chemical components in the water extract of S. glabra (ZJF) were analyzed by UPLC-MS/MS method. The HCoEpiC cell line was used to assess the promotive effect on intestinal proliferation and restitution. RAW264.7 cells were used to assess the in vitro anti-inflammatory effect of ZJF. The 3% DSS-induced colitis model was used to evaluate the in vivo effect of ZJF (4.5 g/kg and 9.0 g/kg). Mesalazine (0.5 g/kg) was used as the positive drug. ELISA, RT-qPCR, Western blot, and multiplex immunohistochemical experiments were used to test gene levels in the colon tissue. The H&E staining method was used to monitor the pathological changes of colon tissue. TUNEL assay kit was used to detect apoptosis of epithelial colonic cells. RESULTS: ZJF could alleviate the DSS-caused colitis in colon tissues, showing a comparative effect to that of the positive drug mesalazine. Mechanism study indicated that ZJF could promote normal colonic HCoEpiC cell proliferation and restitution, inhibit overexpression of pro-inflammatory cytokines, restore the M1/M2 ratio, decrease epithelial colonic cell apoptosis, rescue tight junction protein levels, and modulate IL-17/Notch1/FoxP3 pathway to treat experimental UC. CONCLUSION: Our results indicated that S. glabra can promote intestinal cell restitution, balance immune response, and modulate IL-17/Notch1/FoxP3 pathway to treat experimental UC.

Sinomenine regulates the cholinergic anti-inflammatory pathway to inhibit TLR4/NF-κB pathway and protect the homeostasis in brain and gut in scopolamine-induced Alzheimer's disease mice.

Sinomenine (SIN), an alkaloid extracted from the Chinese herbal medicine Sinomenium acutum, has great potential in anti-inflammatory, immune regulation, analgesic and sedative, and is already a clinical drug for the treatment of rheumatoid arthritis in China. Our previous studies show SIN inhibits inflammation by regulating ɑ7nAChR, a key receptor of cholinergic anti-inflammatory pathway (CAP), which plays an important role in regulating peripheral and central nervous system inflammation. Growing evidence supports the cholinergic dysregulation and inflammatory responses play the key role in the pathogenesis of AD. The intervention effects of SIN on AD by regulating CAP and homeostasis in brain and gut were analyzed for the first time in the present study using scopolamine-induced AD model mice. Behavioral tests were used to assess the cognitive performance. The neurons loss, cholinergic function, inflammation responses, biological barrier function in the mouse brain and intestinal tissues were evaluated through a variety of techniques, and the gut microbiota was detected using 16SrRNA sequencing. The results showed that SIN significantly inhibited the cognitive decline, dysregulation of cholinergic system, peripheral and central inflammation, biological barrier damage as well as intestinal flora disturbance caused by SCOP in mice. More importantly, SIN effectively regulated CAP to suppress the activation of TLR4/NF-κB and protect the homeostasis in brain and gut to alleviate cognitive impairment.

Protective effect of valerian extract capsule (VEC) on ethanol- and indomethacin-induced gastric mucosa injury and ameliorative effect of VEC on gastrointestinal motility disorder.

CONTEXT: Valerian extract capsule (VEC) is an effective Chinese patent medicine used for gastrointestinal (GI) diseases. OBJECTIVE: To investigate the detailed pharmacological activity for VEC clinical effects in GI diseases. MATERIALS AND METHODS: Sprague-Dawley rats were divided into six groups: control, model, and drug-treated (VEC-L, VEC-M, VEC-H, and teprenone). Rats were orally administered VEC (124, 248, 496 mg/kg) and teprenone (21.43 mg/kg) for 3 consecutive days. After 1 h, the five groups (except the control group) were orally given ethanol (10 mL/kg) for 1 h or indomethacin (80 mg/kg) for 7 h. The spasmolytic activity of VEC (0.01-1 mg/mL) on ACh/BaCl2-induced New Zealand rabbit smooth muscle contraction was performed. The C57BL/6 mice carbon propelling test evaluated the effects of VEC (248-992 mg/kg) on intestinal motility in normal and neostigmine/adrenaline-induced mice. RESULTS: Compared with the model group, VEC treatment reduced the gastric lesion index and mucosal damage. Further experiments showed that the pathological ameliorative effect of VEC was accompanied by augmentation of the enzymatic antioxidant system and cytoprotective marker (COX-1, p < 0.01; PGI2 p < 0.05;), along with the alleviation of the levels of MPO (ethanol: 15.56 ± 0.82 vs. 12.15 ± 2.60, p < 0.01; indomethacin: 9.65 ± 3.06 vs. 6.36 ± 2.43, p < 0.05), MDA (ethanol: 1.66 ± 0.44 vs. 0.81 ± 0.58, p < 0.01; indomethacin: 1.71 ± 0.87 vs. 1.09 ± 0.43, p < 0.05), and inflammatory mediators. VEC decreased the high tone induced by ACh/BaCl2 and promoted intestinal transit in normal and neostigmine/adrenaline-induced mice. DISCUSSION AND CONCLUSIONS: VEC showed a potential gastroprotective effect, suggesting that VEC is a promising phytomedicine for the treatment of GI diseases.

Sinomenine inhibits macrophage M1 polarization by downregulating α7nAChR via a feedback pathway of α7nAChR/ERK/Egr-1.

BACKGROUND: Sinomenine (SIN) is an anti-inflammatory drug that has been used for decades in China to treat arthritis. In a previous study, SIN acted on α7 nicotinic acetylcholine receptor (α7nAChR) to inhibit inflammatory responses in macrophages, which indicates a new anti-inflammatory mechanism of SIN. However, the level of α7nAChR was increased in the inflammatory responses and was downregulated by SIN in vitro, so the underlying mechanisms of SIN acting on α7nAChR remain unclear. PURPOSE: To analyze the role of α7nAChR in inflammation and the effect and mechanism of SIN regulation of α7nAChR. METHODS: The effects of SIN on α7nAChR in endotoxemic mice and LPS-stimulated macrophages were observed. Nicotine (Nic) was used as a positive control, and berberine (Ber) was used as a negative control targeting α7nAChR. The antagonists of α7nAChR, α-bungarotoxin (BTX) and mecamylamine (Me), were used to block α7nAChR. In RAW264.7 macrophage cells in vitro, α7nAChR short hairpin RNA (shRNA) was used to knock down α7nAChR. Macrophage polarization was analyzed by the detection of TNF-α, IL-6, iNOS, IL-10, Arg-1, and Fizz1. U0126 was used to block ERK phosphorylation. The cytokines α7nAChR, ERK1/2, p-ERK1/2 and Egr-1 were detected. RESULTS: SIN decreased the levels of TNF-α, IL-6 and the expression of α7nAChR increased by LPS in endotoxemic mice. The above effects of SIN were attenuated by BTX. In the α7nAChR shRNA transfected RAW264.7 cells, compared with the control, α7nAChR was knocked down, and M1 phenotype markers (including TNF-α, IL-6, and iNOS) were significantly downregulated, whereas M2 phenotype markers (including IL-10, Arg-1, and Fizz1) were significantly upregulated when stimulated by LPS. SIN inhibited the expression of p-ERK1/2 and the transcription factor Egr-1 induced by LPS in RAW264.7 cells, and the above effects of SIN were attenuated by BTX. The expression of α7nAChR was suppressed by U0126, which lessened the expression of p-ERK1/2 and Egr-1. CONCLUSIONS: SIN acts on α7nAChR to inhibit inflammatory responses and downregulates high expression of α7nAChR in vivo and in vitro. The increase of α7nAChR expression is correlated with inflammatory responses and participates in macrophage M1 polarization. SIN downregulates α7nAChR via a feedback pathway of α7nAChR/ERK/Egr-1, which contributes to inhibiting macrophage M1 polarization and inflammatory responses.

Glycyrrhetinic acid reverses antibiotic-induced intestinal epithelial injury through RNA-binding protein human antigen R (HuR).

BACKGROUND: The application/abuse of antibiotics can cause antibiotic-induced intestinal injury (AIJ), a typical clinical issue that disturbs intestinal homeostasis. However, the underlying post-transcriptional mechanism of AIJ remains unknown. Glycyrrhetinic acid (GA) is one of the main components of Glycyrrhiza uralensis Fisch. and Glycyrrhiza inflata Batalin (Fabaceae), and findings of our previous study showed that GA can maintain intestinal homeostasis post-transcriptionally through the RNA-binding protein human antigen R (HuR). PURPOSE: This study aimed to elucidate the role of HuR in AIJ and the protective effects of GA on AIJ. STUDY DESIGN AND METHODS: Clindamycin hydrochloride was used to clarify the effect of the antibiotic on the intestinal epithelium. Intestinal epithelium cell-6 (IEC-6) and Caco2 cells were used to demonstrate the in vitro effects of the antibiotic and GA on intestinal cells. HuR plasmid and siRNA were used to overexpress or silence HuR in vitro. SD rats were induced by using clindamycin hydrochloride capsules (250 mg/kg i.g.) for 7 consecutive days to construct the in vivo AIJ model. Rats of the AIJ model group were administrated GA (10 and 20 mg/kg i.g.) for 7 days, and subsequently, the protective effect of GA on the intestinal epithelium was evaluated. RESULTS: In vitro results showed that the antibiotic (150-500 µM) suppressed proliferation, induced a delay in restitution after wounding, and caused cell cycle arrest at the G0/G1 phase in IEC-6 and Caco-2 cells. Moreover, the expression levels of HuR and its downstream gene, occludin and cyclin D1, decreased after treatment with the antibiotic (500 µM). Overexpression of HuR and GA (10 and 20 µM) reversed the antibiotic-induced inhibition of proliferation and G0/G1 phase arrest, and the antibiotic-induced decrease in HuR, occludin, and cyclin D1 expression was reversed after GA treatment (10 and 20 µM) in IEC-6 cells. In vivo results revealed the antibiotic-induced epithelial injury of both the small intestines (shortened and spared mucosa) and the large intestines (injured/deformed glands, reduced number of cup cells, and evident inflammatory cell infiltration), all of which were ameliorated after GA treatment (10 and 20 µM). CONCLUSION: Antibiotics induce intestinal epithelial injury through HuR, and GA can exert a protective effect on AIJ by restoring HuR levels.

Qiangji Jianli Decoction Alleviates Hydrogen Peroxide-Induced Mitochondrial Dysfunction via Regulating Mitochondrial Dynamics and Biogenesis in L6 Myoblasts.

Oxidative stress can cause the excessive generation of reactive oxygen species (ROS) and has various adverse effects on muscular mitochondria. Qiangji Jianli decoction (QJJLD) is an effective traditional Chinese medicine (TCM) that is widely applied to improve muscle weakness, and it has active constituents that prevent mitochondrial dysfunction. To investigate the protective mechanism of QJJLD against hydrogen peroxide- (H2O2-) mediated mitochondrial dysfunction in L6 myoblasts. Cell viability was determined with MTT assay. Mitochondrial ultrastructure was detected by transmission electron microscope (TEM). ROS and mitochondrial membrane potential (MMP) were analyzed by fluorescence microscope and flow cytometry. The superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) activity, and malondialdehyde (MDA) level were determined by WST-1, TBA, and DTNB methods, respectively. The mRNA and protein levels were measured by quantitative real-time PCR (qRT-PCR) and Western blot. The cell viability was decreased, and the cellular ROS level was increased when L6 myoblasts were exposed to H2O2. After treatment with QJJLD-containing serum, the SOD and GSH-Px activities were increased. MDA level was decreased concurrently. ROS level was decreased while respiratory chain complex activity and ATP content were increased in L6 myoblasts. MMP loss was attenuated. Mitochondrial ultrastructure was also improved. Simultaneously, the protein expressions of p-AMPK, PGC-1α, NRF1, and TFAM were upregulated. The mRNA and protein expressions of Mfn1/2 and Opa1 were also upregulated while Drp1 and Fis1 were downregulated. These results suggest that QJJLD may alleviate mitochondrial dysfunction through the regulation of mitochondrial dynamics and biogenesis, the inhibition of ROS generation, and the promotion of mitochondrial energy metabolism.

Kuijieling decoction suppresses NLRP3-Mediated pyroptosis to alleviate inflammation and experimental colitis in vivo and in vitro.

ETHNOPHARMACOLOGICAL RELEVANCE: Ulcerative colitis (UC) is an autoimmune disease. Although the mortality rate of UC is not very high, it has a considerable morbidity rate and an unsatisfactory cure rate. Without effective treatment, UC is likely to develop into colon cancer. Kuijieling (KJL) is an effective empirical formula to treat UC in the clinical setting, and it has been proven to have curative effects against UC. AIM OF THE STUDY: In a previous study, we demonstrated that KJL could suppress NOD-like receptor protein 3 (NLRP3) to reduce inflammatory cytokines and alleviate UC. In this study, we investigated the mechanism of KJL in more detail, from the perspective of pyroptosis. MATERIALS AND METHODS: We established a dextran sulfate sodium-induced UC mouse model and RAW264.7 cells to measure different indicators with different experimental methods. The efficiency of KJL was evaluated by measuring the length and unit weight of mouse colons, and assessment of pathological injury was performed using HE staining. We detected different expression levels of NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), caspase-1, gasdermin-D C-terminal domain (GSDMD-C), gasdermin-D N-terminal domain (GSDMD-N), IL-1ß, and IL-18 in colon tissues and cells using RT-qPCR and western blotting. Immunohistochemistry was used for tissues and immunofluorescence for cells to confirm protein expression. IL-1ß and IL-18 were measured with enzyme-linked immunosorbent assay in serum, tissue, and cell culture supernatant. MiR-223 was detected using RT-qPCR. RESULTS: After administration of KJL suspension, colon damage in KJL groups was milder than in model groups. ASC, caspase-1, IL-1ß, and IL-18 mRNA levels in colon tissue were decreased to different degrees in the KJL groups. Protein expression of NLRP3, caspase-1, GSDMD-N, IL-1ß, and IL-18 in vivo decreased significantly in the KJL groups. In addition, Mir-223 level decreased in colon tissue of the KJL groups. In vitro, NLRP3, ASC, caspase-1, GSDMD-N, IL-1ß, and IL-18 levels decreased to varying degrees, at both mRNA and protein levels. Mir-223 was lower in the KJL groups than in the model group. Furthermore, KJL was shown to regulate the level of miR-223, which returned to normal after its expression was inhibited or promoted, and the levels of associated indicators also returned to normal after transfection. CONCLUSIONS: KJL is able to inhibit pyroptosis to alleviate UC, but these suppression effects were not mediated through miR-223 regulation.

Ophiopogonin D' induces RIPK1­dependent necroptosis in androgen­dependent LNCaP prostate cancer cells.

Ophiopogonin D' (OPD') is a natural compound extracted from Ophiopogon japonicus, which is a plant used in traditional Chinese medicine. Our previous study has indicated that OPD' exhibits antitumor activity against androgen­independent prostate cancer (PCa), but the effects and the underlying molecular mechanism of action of OPD' in androgen­dependent PCa were unclear. In the present study, OPD' induced significant necroptosis in androgen­dependent LNCaP cancer cells by activating receptor­interacting serine/threonine­protein kinase 1 (RIPK1). Exposure to OPD' also increased Fas ligand (FasL)­dependent RIPK1 protein expression. The OPD'­induced necroptosis was inhibited by a RIPK1 inhibitor necrostatin­1, further supporting a role for RIPK1 in the effects of OPD´. The antitumor effects of OPD' were also inhibited by a mixed lineage kinase domain­like protein (MLKL) inhibitor necrosulfonamide. Following treatment with inhibitors of RIPK1 and MLKL, the effects of OPD' on LNCaP cells were inhibited in an additive manner. In addition, co­immunoprecipitation assays demonstrated that OPD' induced RIPK3 upregulation, leading to the assembly of a RIPK3­MLKL complex, which was independent of RIPK1. Furthermore, OPD' increased the expression of Fas­associated death domain, which is required to induce necroptosis in LNCaP cells. OPD' also regulated the expression levels of FasL, androgen receptor and prostate­specific antigen in a RIPK1­dependent manner. These results suggested that OPD' may exhibit potential as an anti­PCa agent by inducing RIPK1­ and MLKL­dependent necroptosis.

Patchouli alcohol activates PXR and suppresses the NF-κB-mediated intestinal inflammatory.

ETHNOPHARMACOLOGICAL RELEVANCE: The pregnane-X-receptor (PXR) is involved in inflammatory bowel disease (IBD). Patchouli alcohol (PA) has anti-inflammatory effects; however, the effect of PA on IBD pathogenesis remains largely unknown. AIM OF THE STUDY: The aim of the present study was to investigate the anti-inflammatory effect of PA, primarily focused on crosstalk between PA-mediated PXR activation and NF-κB inhibition. MATERIALS AND METHODS: We evaluated the anti-inflammatory effect of PA with respect to PXR/NF-κB signalling using in vitro and in vivo models. In vitro, PA, identified as a PXR agonist, was evaluated by hPXR transactivation assays and through assessing for CYP3A4 expression and activity. NF-κB inhibition was analysed based on NF-κB luciferase assays, NF-κB-mediated pro-inflammatory gene expression, and NF-κB nuclear translocation after activation of PXR by PA. In vivo, colonic mPXR and NF-κB signalling were analysed to assess PA-mediated the protective effect against dextran sulphate sodium (DSS)-induced colitis. Furthermore, pharmacological inhibition of PXR was further evaluated by examining PA protection against DSS-induced colitis. RESULTS: PA induced CYP3A4 expression and activity via an hPXR-dependent mechanism. PA-mediated PXR activation attenuated inflammation by inhibiting NF-κB activity and nuclear translocation. The anti-inflammatory effect of PA on NF-κB was abolished by PXR knockdown. PA prevented DSS-induced inflammation by regulating PXR/NF-κB signalling, whereas pharmacological PXR inhibition abated PA-mediated suppressive effects on NF-κB inflammation signalling. CONCLUSIONS: PA activates PXR signalling and suppresses NF-κB signalling, consequently causing amelioration of inflammation. Our results highlight the importance of PXR-NF-κB crosstalk in colitis and suggest a novel therapeutic reagent.

Kuijieling-Containing Serum Regulates Th17 and Treg Cell Differentiation by Inhibiting STAT3 Signaling In Vitro.

OBJECT: To investigate the effect of Kuijieling (KJL) on the balance between T helper 17 (Th17) and regulatory T (Treg) cells in peripheral blood mononuclear cells (PBMC) in vitro and explore the underlying mechanism. MATERIALS AND METHODS: PBMCs isolated from rats were stimulated with transforming growth factor-ß, interleukin (IL)-6, and IL-23 to induce the imbalance of Th17 and Treg cells and were treated with 10, 5, or 2.5% KJL-containing serum. The proportion of Th17 or Treg cells in CD4+ T cells was analyzed by flow cytometry, the concentrations of IL-17, IL-21, and IL-10 were assayed by ELISA, mRNA expressions of retinoic acid-related orphan receptor γt (RORγt), forkhead box protein 3 (Foxp3), and signal transducer and activator of transcription 3 (STAT3) were quantified by PCR, and phosphorylated STAT3 (p-STAT3) was analyzed by flow cytometry. RESULTS: KJL-containing serum decreased the proportion of Th17 cells and increased the proportion of Treg cells in CD4+ T cells, decreased the concentration of IL-17 and IL-21, enhanced the level of IL-10 in the cell culture supernatant, promoted the expression of Foxp3, and inhibited the levels of RORγt, STAT3, and p-STAT3. CONCLUSION: KJL suppresses the STAT3 pathway to remedy the imbalance between Th17 and Treg cells.

Extract of Ilex rotunda Thunb alleviates experimental colitis-associated cancer via suppressing inflammation-induced miR-31-5p/YAP overexpression.

BACKGROUND: Ilex rotunda Thunb is a traditional medicine used in China treating colitis clinically. Triterpenoids is one of its main components. However, the detailed pharmacological activity and the component responsible for its clinical effects are still elusive. PURPOSE: To test the in vivo colitis-associated cancer (CAC) preventive effect of the water fraction extracted from the roots of I. rotunda, and to evaluate its microRNA (miRNA)-related mechanism. STUDY DESIGN AND METHODS: Male or female C57BL/6 mice (12 weeks of age) were used to construct the azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced CAC. 12.5 mg/kg and 25.0 mg/kg of the standardized water extract of I. rotunda (WIR), being equal to 4.29 and 8.58 g of the raw medicine respectively, were adopted to treat the AOM/DSS-induced CAC from the fourth week and continued for 5 weeks. Mice were killed two weeks after the end of the last round of DSS by cervical dislocation. RESULTS: The chemical analysis of WIR revealed the presence of 21 compounds. The syringing and caffeic acid (1-hydroxyl-4-O-ß-D-glucopyranosylprenyl)-ester are the main components of WIR, counting for 8.27% and 5.71% of the water extract respectively. The levels of miR-31-5p were up-regulated in both thp1 and Caco2 cells (p < 0.05) stimulated by either IL-6 or TNF-α, and WIR could restore miR-31-5p levels in the IL-6/TNF-α-stimulated thp-1 and Caco2 cells. Furthermore, WIR decreased TNF-α and IL-6 levels in PMA-differentiated thp-1 cells stimulated by LPS via NF-κB pathway (p < 0.05), suggesting that WIR could restore miR-31-5p expression via down-regulating IL-6 and TNF-α levels. In vivo study showed that oral administration of WIR (25 mg/kg) produced a significant inhibition on the atypical hyperplasia, as well as the release and the expression of IL-6 and TNF-α in the colon tissue. The in vivo transcription of other pro-inflammatory mediators such as iNOS, IL-11, and IL-17A were also attenuated by WIR administration (25 mg/kg, p < 0.05). Meanwhile, WIR (25 mg/kg) restored the miR-31-5p level which was up-regulated in the CAC model group, and ectopic expressions of the miR-31-5p down-stream LATS2 and YAP genes in the hippo pathway were also modulated by the WIR (25 mg/kg) treatment. CONCLUSION: The present study suggests that WIR exerts intestinal anti-inflammatory and CAC preventive effects in an experimental CAC mouse model. The CAC preventive effect can be attributed to the suppression of hippo pathway activated by the inflammatory cytokines, indicating that WIR can be potentially used as an herbal product for CAC prevention. Therefore, there is an emergent need for further evaluation of the main components in WIR to determine the definite bioactive component responsible for the CAC preventive activity.

HBD Inhibits the Development of Colitis-Associated Cancer in Mice via the IL-6R/STAT3 Signaling Pathway.

Colitis-associated cancer (CAC) is a malignant disease of the colon that is caused by recurrent episodes of chronic intestinal inflammation. Huangqi Baizhu decoction (HBD) is a classic prescription comprised of Radix Astragali and Rhizoma Atractylodis, which are usually used to treat digestive conditions, such as peptic ulcers, colitis, or colorectal carcinoma in clinics. HBD is well known for "tonifying qi and spleen" based on the theories of traditional Chinese medicine, and has the preponderant effect of alleviating chronic intestinal mucosa damage associated with disease. However, the underlying mechanism behind this is still unknown. In the current study, we employed the AOM/DSS mouse model to analyze the effects of HBD on the development of inflammation in colonic carcinoma. The in vivo study showed that HBD could significantly reduce the mortality of mice and control the incidence and size of colonic tumors by inhibiting the IL-6/STAT3 signaling pathway. In vitro, Astragaloside and Atractylenolide (CAA), the main components of HBD, inhibited the proliferation of HCT-116 cells as determined by an MTT assay. Furthermore, CAA notably suppressed the protein expression of IL-6R, STAT3, Survivin, and Cyclin D1 induced by IL-6 in HCT-116 and RAW264.7 cells. These results suggested that HBD exhibits anti-inflammatory and anti-proliferative effects, inhibiting the development of CAC in mice.

Evaluation of the anticancer and anti-metastasis effects of novel synthetic sodium channel blockers in prostate cancer cells in vitro and in vivo.

BACKGROUND: Voltage-gated sodium channels (VGSCs) are involved in several cellular processes related to cancer cell growth and metastasis, including adhesion, proliferation, apoptosis, migration, and invasion. We here in investigated the effects of S0154 and S0161, two novel synthetic sodium channel blockers (SCBs), on human prostate cancer cells (PC3, DU145, and LnCaP) and a prostate cancer xenograft model. METHODS: The MTT assay was used to assess the anticancer effects of SCBs in PC3, DU145, and LnCaP cells. Sodium indicator and glucose uptake assays were used to determine the effects of S0154 and S0161 in PC3 cells. The impact of these SCBs on the proliferation, cell cycle, apoptosis, migration, and invasion of PC3 cells were determined using a CFDA-SE cell proliferation assay, cell cycle assay, annexin V-FITC apoptosis assay, transwell cell invasion assay, and wound-healing assay, respectively. The protein expression levels of Nav1.6, Nav1.7, CDK1, cyclin B1, MMP2, MMP9 in PC3 cells were analysis by Western blotting. The in vivo anticancer activity was evaluated using a PC3 xenograft model in nude mice. RESULTS: S0154 and S0161 both showed anticancer and anti-metastatic effects against prostate cancer cells and significantly inhibited cell viability, with IC50 values in the range of 10.51-26.60 µmol/L (S0154) and 5.07-11.92 µmol/L (S0161). Both compounds also increased the intracellular level of sodium, inhibited the protein expression of two α subunits of VGSCs (Nav1.6 and Nav1.7), and caused G2/M phase cell cycle arrest, with no or minor effects on cell apoptosis. Concentrations of 5 and 10 µmol/L of S0154 and S0161 significantly decreased the glucose uptake of PC3 cells. The compounds also inhibited the proliferation of PC3 cells and decreased their invasion in transwell assays. Furthermore, S0161 exerted antitumor activity in an in vivo PC3 xenograft model in nude mice, inhibiting the growth of the tumors by about 51% compared to the control group. CONCLUSIONS: These results suggest that S0154 and S0161 have anticancer and anti-metastasis effects in prostate cancer cells both in vitro and in vivo, supporting their further development as potential therapeutic agents for prostate cancer.

Kuijieling regulates the differentiation of Treg and Th17 cells to ameliorate experimental colitis in rats.

BACKGROUND: Regulatory T (Treg) cells and T helper 17 (Th17) cells play crucial roles in ulcerative colitis (UC). Kuijieling (KJL) is an effective Chinese medicine formula for treating UC in clinic. Kuijieling has shown remedy effect on the imbalance between Treg and Th17 cells. This study aimed to further reveal the exact underlying mechanism of how Kuijieling regulates the differentiation of Treg and Th17 cells in the treatment of UC. METHODS: Colitis was induced by trinitrobenzene sulfonic acid in rats and treated by KJL. Pathological injury was evaluated by HE staining and pathological score. Transforming growth factor-ß1 (TGF-ß1), interleukin(IL)-2, IL-6, IL-10, IL-17, IL-23 and IL-21 in plasma were assayed by ELISA. Forkhead box P3 (Foxp3), signal transducer and activator of transcription (STAT) 5 expressed in colon mucosa were measured by western blot. Immunohistochemistry was employed for quantifying retinoic acid-related orphan receptor γt (RORγt) and STAT3 in colon. RT-PCR was used to analyze the expression of IL-2, IL-17, IL-23, IL-21 mRNA in colon. RESULTS: After the administration of KJL, pathological injury in colon mucosa was reduced and histological score was decreased, transforming growth factor-ß1 (TGF-ß1), interleukin(IL)-2, IL-10 in blood and Foxp3, STAT5, IL-2 in colon increased significantly, IL-6, IL-23, IL-17, IL-21 in blood and RORγt, STAT3, IL-23, IL-17, IL-21 in colon decreased. Our result showed that KJL regulates the related cytokines and transcription factors to promote Treg cells and suppress Th17 cells. CONCLUSION: KJL restores the balance between Treg and Th17 cells through regulating the differentiation of them, therefore contributes to the treatment of UC.

Ophiopogonin D', a Natural Product From Radix Ophiopogonis, Induces in Vitro and in Vivo RIPK1-Dependent and Caspase-Independent Apoptotic Death in Androgen-Independent Human Prostate Cancer Cells.

Objective: The purpose of this study was to evaluate the anticancer effects of Ophiopogonin D' (OPD', a natural product extracted from a traditional Chinese medicine (Radix Ophiopogonis) against androgen-independent prostate cancer cells and to explore the underlying molecular mechanism(s) of action. Methods: The CCK-8 assay was used to assess the viability of prostate cancer cells. The cell morphology was examined by an ultrastructural analysis via transmission electron microscopy. Cells in apoptosis (early and late stages) were detected using an Annexin V-FITC/propidium iodide kit with a FACSCaliber flow cytometer. JC-1, a cationic lipophilic probe, was employed to measure the mitochondrial membrane potential (MMP) of PC3 cells. Changes in the protein expression of RIPK1, C-RIPK1, caspase 8, cleaved-caspase 8, Bim, Bid, caspase 10, and cleaved-caspase 10 were evaluated by Western blotting. The mRNA expression of Bim was examined by quantitative real-time reverse transcription polymerase chain reaction. Z-VAD-FMK (a caspase inhibitor) and necrostatin-1 (a specific inhibitor of RIPK1) were utilized to determine whether the cell death was mediated by RIPK1 or caspases. PC3 and DU145 xenograft models in BALB/c nude mice were used to evaluate the anticancer activity of OPD' in vivo. Results: OPD' was shown to exert potent anti-tumor activity against PC3 cells. It induced apoptosis via a RIPK1-related pathway, increased the protein expression levels of RIPK1 and Bim, and decreased the levels of cleaved-RIPK1, caspase 8, cleaved-caspase 8, Bid, caspase 10, and cleaved-caspase 10. OPD' also increased the mRNA expression of Bim. The protein expression of Bim was decreased when cells were pre-treated with necrostatin-1. Treatment with OPD' inhibited the growth of PC3 and DU145 xenograft tumors in BALB/c nude mice. Conclusion: OPD' significantly inhibited the in vitro and in vivo growth of prostate cells via RIPK1, suggesting that OPD' may be developed as a potential anti-prostate cancer agent.

Vitamin D: preventive and therapeutic potential in Parkinson's disease.

Vitamin D is one of the important nuclear steroid transcription regulators that controls transcriptions of a large number of genes. Vitamin D supplement is commonly recommended for the elderly to prevent bone diseases. Amounting new evidence has indicated that vitamin D plays a crucial role in brain development, brain function regulation and neuroprotection. Parkinson's disease (PD) is a degenerative disorder commonly seen in the elderly, characterized by movement disorders including tremor, akinesia, and loss of postural reflexes. The motor symptoms largely result from the continued death of dopaminergic neurons in the substantia nigra, despite use of current therapeutic interventions. The cause and mechanism of neuron death is currently unknown. Vitamin D deficiency is common in patients with PD suggesting its preventive and therapeutic potential. Vitamin D may exert protective and neurotropic effects directly at cellular level, e.g. protection of dopamine system, and/or by regulating gene expression. This review summarizes the epidemiological, genetic and translational evidence implicating vitamin D as a candidate for prevention and treatment for PD.

[Effect of total saponins from Panax notoginseng on expession of melatonin receptor mRNA in gastric mucosa of stress rats].

OBJECTIVE: To observe the effect of total saponins from Panax notoginseng (PNS) on melatonin receptor (MR) expression and its content in gastric mucosa of stress rats. METHODS: Water-immersion restraint stress (WRS) was used to induce stress rat models. RevertAid First Strand cDNA Synthesis Kit was used for the reverse transcription of the total RNA from gastric mucosa of stress rats. The PCR product was analyzed by agarose gel electrophoresis firstly and then by KODAK 1 D Image system. The expression level of melatonin 1 (MT1) receptor mRNA was calculated with the optimal density of ratio of MT, receptor and GAPDH. RESULTS: Six hours after WRS, there were 360 bp positive bands in RT-PCR product of MT1. The relative content of MR was lower in the model group than that in the normal group. MR content in PNS group and ranitidine group were both higher than that in the normal group and the model group (P < 0.05 or P < 0.01). CONCLUSION: There is mRNA expression of MT1 receptor in gastric mucosa of stress rats indeed. PNS has protective effect on gastric mucosa of stress rats by up-regulating MR expression, and its protective mechanism may be related to promoting melatonin secretion or increasing the binding capacity of melatonin to its receptor.

[Effect of Kuijieling decoction on proteins expression of IKK-alpha in colonic mucosa of ulcerative colitis model rats].

OBJECTIVE: To explore the changes of protein expression of IKK-alpha as well as the effects of Kuijieling Decoction (KD) on colonic mucosa of ulcerative colitis (UC) model rats. METHODS: UC model rats were induced by TNBS. The rats were randomly divided into six groups: normal control (NC) group, model control (MC) group, Kuijieling low dose (KLD), middle dose (KMD) group, high dose (KHD) group and SASP group. After 10-days' treatment the rats were killed to get their colonic tissues. The positive rate of IKK-alpha expression was detected by immunohistochemical (IHC). RESULTS: The positive rate of IKK-alpha in MC group was significantly higher than that in NC group (P < 0.01). The positive rate of IKK-alpha in KMD group was significantly lower than that in MC group (P < 0.05). The positive rate of IKK-alpha in KHD and SASP group were significantly lower than that in MC group (P < 0.01). CONCLUSION: IKK-alpha may be involved in the pathogenesis of UC, and KD can inhibit positive rate of IKK-alpha in colonic mucosa of UC model rats induced by TNBS. The inhibitory effects of KD on UC may be associated with this.

Effects of Dangshen root extract serum on intracellular free calcium in gastric parietal cells of dissociated rats.

OBJECTIVE: To study the effects of Dangshen root extract on intracellular free calcium concentration [Ca2+] i of parietal cells. METHODS: The Dangshen-containing serum was obtained from rat blood after a continuously five days' feeding with Dangshen root extract of three doses and parietal cells were isolated from Sprague-Dawley rats; [Ca2+] i in single cells was measured by confocal microscope loaded with Fluo3-AM as fluorensent indicator; The change of [Ca2+] i was represented by fluorensent intensity (FI). RESULTS: There were differences in the FI of [Ca2+] i increased by gastrin both between high and control group and between middle and control group (P < 0.05). But no difference was found between low and control groups (P > 0.05). And time to peak of FI, it was not found any difference between any two of the groups. CONCLUSION: Dangshen-containing serum can inhibit the intracellular [Ca2+] i increase induced by gastrin in a dose-dependent manner and it may be one of the mechanisms of its reduction on acid secretion which has a close relation with the formation of ulcerous diseases.