IL-6 Inhibitory Compounds from the Aerial Parts of Piper attenuatum and Their Anticancer Activities on Ovarian Cancer Cell Lines.

Piper attenuatum Buch-Ham, a perennial woody vine belonging to the Piperaceae family, is traditionally used in Southeast Asia for treating various ailments such as malaria, headache, and hepatitis. This study described the isolation and identification of three new compounds, piperamides I-III (1-3), which belong to the maleimide-type alkaloid skeletons, along with fifteen known compounds (4-18) from the methanol extract of the aerial parts of P. attnuatum. Their chemical structures were elucidated using spectroscopic methods (UV, IR, ESI-Q-TOF-MS, and 1D/2D NMR). All the isolates were evaluated for their ability to inhibit IL-6 activity in the human embryonic kidney-Blue™ IL-6 cell line and their cytotoxic activity against ovarian cancer cell lines (SKOV3/SKOV3-TR) and chemotherapy-resistant variants (cisplatin-resistant A2780/paclitaxel-resistant SKOV3). The compounds 3, 4, 11, 12, 17, and 18 exhibited IL-6 inhibition comparable to that of the positive control bazedoxifene. Notably, compound 12 displayed the most potent anticancer effect against all the tested cancer cell lines. These findings highlight the importance of researching the diverse activities of both known and newly discovered natural products to fully unlock their potential therapeutic benefits.

A derivative of 3-(1,3-diarylallylidene)oxindoles inhibits dextran sulfate sodium-induced colitis in mice.

BACKGROUND: IA-0130 is a derivative of 3-(1,3-diarylallylidene)oxindoles, which is a selective estrogen receptor modulator (SERM). A previous study demonstrated that SERM exhibits anti-inflammatory effects on colitis by promoting the anti-inflammatory phenotype of monocytes in murine colitis. However, the therapeutic effects of oxindole on colitis remain unknown. Therefore, we evaluated the efficacy of IA-0130 on dextran sulfate sodium (DSS)-induced mouse colitis. METHODS: The DSS-induced colitis mouse model was established by administration of 2.5% DSS for 5 days. Mice were orally administered with IA-0130 (0.01 mg/kg or 0.1 mg/kg) or cyclosporin A (CsA; 30 mg/kg). Body weight, disease activity index score and colon length of mice were calculated and histological features of mouse colonic tissues were analyzed using hematoxylin and eosin staining. The expression of inflammatory cytokines and tight junction (TJ) proteins were analyzed using quantitative real-time PCR and enzyme-linked immunosorbent assay. The expression of interleukin-6 (IL-6) signaling molecules in colonic tissues were investigated using Western blotting and immunohistochemistry (IHC). RESULTS: IA-0130 (0.1 mg/kg) and CsA (30 mg/kg) prevented colitis symptom, including weight loss, bleeding, colon shortening, and expression of pro-inflammatory cytokines in colon tissues. IA-0130 treatment regulated the mouse intestinal barrier permeability and inhibited abnormal TJ protein expression. IA-0130 down-regulated IL-6 expression and prevented the phosphorylation of signaling molecules in colonic tissues. CONCLUSIONS: This study demonstrated that IA-0130 suppressed colitis progression by inhibiting the gp130 signaling pathway and expression of pro-inflammatory cytokines, and maintaining TJ integrity.

Geraniin Alleviates Inflammation in Caco-2 Cells and Dextran Sulfate Sodium-Induced Colitis Mice by Targeting IL-1ß.

IL-1ß is an important cytokine implicated in the progression of inflammatory bowel disease (IBD) and intestinal barrier dysfunction. The polyphenolic compound, geraniin, possesses bioactive properties, such as antitumor, antioxidant, anti-inflammatory, antihypertensive, and antiviral activities; however, its IL-1ß-targeted anticolitis activity remains unclear. Here, we evaluated the inhibitory effect of geraniin in IL-1ß-stimulated Caco-2 cells and a dextran sulfate sodium (DSS)-induced colitis mouse model. Geraniin blocked the interaction between IL-1ß and IL-1R by directly binding to IL-1ß and inhibited the IL-1ß activity. It suppressed IL-1ß-induced intestinal tight junction damage in human Caco-2 cells by inhibiting IL-1ß-mediated MAPK, NF-kB, and MLC activation. Moreover, geraniin administration effectively reduced colitis symptoms and attenuated intestinal barrier injury in mice by suppressing elevated intestinal permeability and restoring tight junction protein expression through the inhibition of MAPK, NF-kB, and MLC activation. Thus, geraniin exhibits anti-IL-1ß activity and anticolitis effect by hindering the IL-1ß and IL-1R interaction and may be a promising therapeutic anti-IL-1ß agent for IBD treatment.

Celastrol regulates psoriatic inflammation and autophagy by targeting IL-17A.

Anti-IL-17A antibodies, such as secukinumab and ixekizumab, are effective proinflammatory cytokine inhibitors for autoimmune disorders, including psoriasis. However, anti-IL-17A small molecule treatments are yet to be commercialized. Celastrol, a natural compound extracted from the roots of traditional Chinese medicinal plants, has anti-inflammatory and antioxidant properties. However, the binding of celastrol to IL-17A and the associated anti-inflammatory mechanisms remain unclear. This study investigated whether celastrol could directly bind to IL-17A and regulate inflammation in psoriatic in vitro and in vivo models. The results showed that celastrol directly binds to IL-17A and inhibits its downstream signaling, including the NF-kB and MAPK pathways. Interestingly, celastrol restored autophagy dysfunction and reduced proinflammatory cytokine secretion in keratinocytes. In addition, celastrol increased autophagy in the epidermis of a mouse model of psoriasis. Celastrol decreased Th17 cell populations and proinflammatory cytokine levels in mice. Thus, IL-17A-targeting celastrol reduced inflammation by rescuing impaired autophagy in in vitro and in vivo models of psoriasis, demonstrating its potential as a substitute for anti-IL-17A antibodies for treating psoriasis.

Celastrol, which targets IL-2/CD25 binding inhibition, induces T cell-mediated antitumor activity in melanoma.

Interleukin-2 (IL-2) induces contrasting immune responses depending on its binding receptor subunit; thus, selective receptor binding is considered a key challenge in cancer therapeutic strategies. In this study, we aimed to investigate the inhibition of IL-2 action and antitumor activity of celastrol (CEL), a compound identified in a screen for IL-2/CD25 binding inhibitors, and to elucidate the underlying role of CEL in immune cells. We found that CEL selectively impairs the binding of IL-2 and CD25 and directly binds to IL-2 but not to CD25. CEL significantly suppressed the proliferation and signaling of IL-2-dependent murine T cells and interfered with IL-2-responsive STAT5 phosphorylation in IL-2 reporter cells and human PBMCs. After confirming the impact of CEL on IL-2, we evaluated its antitumor activity in C57BL/6 mice bearing B16F10 tumors and found that CEL significantly inhibited tumor growth by increasing CD8+ T cells. We also found that CEL did not inhibit tumor growth in T cell-deficient BALB/c nude mice, suggesting that its activity was mediated by the T-cell response. Moreover, combination therapy with low-dose CEL and a TNFR2 antagonist synergistically improved the therapeutic efficacy of the individual monotherapies by increasing the ratio of intratumoral CD8/Treg cells and suppressing Foxp3 expression. These findings suggest that CEL, which inhibits CD25 binding by targeting IL-2, exerts antitumor activity by mediating the T-cell response and could be a promising candidate for combination therapy in cancer immunotherapy against melanoma.

Aberrant glucose metabolism underlies impaired macrophage differentiation in glycogen storage disease type Ib.

Glycogen storage disease type Ib (GSD-Ib) is an autosomal recessive disorder caused by a deficiency in the glucose-6-phosphate (G6P) transporter (G6PT) that is responsible for transporting G6P into the endoplasmic reticulum. GSD-Ib is characterized by disturbances in glucose homeostasis, neutropenia, and neutrophil dysfunction. Although some studies have explored neutrophils abnormalities in GSD-Ib, investigations regarding monocytes/macrophages remain limited so far. In this study, we examined the impact of G6PT deficiency on monocyte-to-macrophage differentiation using bone marrow-derived monocytes from G6pt-/- mice as well as G6PT-deficient human THP-1 monocytes. Our findings revealed that G6PT-deficient monocytes exhibited immature differentiation into macrophages. Notably, the impaired differentiation observed in G6PT-deficient monocytes seemed to be associated with abnormal glucose metabolism, characterized by enhanced glucose consumption through glycolysis, even under quiescent conditions with oxidative phosphorylation. Furthermore, we observed a reduced secretion of inflammatory cytokines in G6PT-deficient THP-1 monocytes during the inflammatory response, despite their elevated glucose consumption. In conclusion, this study sheds light on the significance of G6PT in monocyte-to-macrophage differentiation and underscores its importance in maintaining glucose homeostasis and supporting immune response in GSD-Ib. These findings may contribute to a better understanding of the pathogenesis of GSD-Ib and potentially pave the way for the development of targeted therapeutic interventions.

IL-17A-targeting fenofibrate attenuates inflammation in psoriasis by inducing autophagy.

IL-17A is a critical pro-inflammatory cytokine in autoimmune diseases such as psoriasis. Targeting of IL-17A is an effective strategy to treat patients with autoimmune diseases; however, relevant small molecule therapeutics have not yet been developed. Here, the small molecule drug fenofibrate was validated as an inhibitor of IL-17A through ELISA and surface plasmon resonance (SPR) assays. We further confirmed that fenofibrate blocked IL-17A signalings including the mitogen-activated protein kinase (MAPK) and NF-κB signaling pathways, in IL-17A-treated HaCaT cells, HEKa (human primary epidermal keratinocytes) and imiquimod (IMQ)-induced psoriasis mouse model. Fenofibrate attenuated systemic inflammation by suppressing Th17 populations and inflammatory cytokines, such as IL-1ß, IL-6, IL-17A, and tumor necrosis factor (TNF). Surprisingly, fenofibrate upregulated LC3 and p62 in the psoriatic mouse group. The autophagy changes were caused by ULK1 pathway in hIL-17A-treated HaCaT and HEKa. In addition, the enhancement of autophagy by fenofibrate exerted anti-inflammatory effects, as demonstrated by the suppression of IL-6 and IL-8 in the IL-17A-treated keratinocytes. Thus, IL-17A-targeting fenofibrate can be a potential therapeutic for psoriasis and other autoimmune diseases via regulating autophagy.

LncRNA SNHG4 Modulates EMT Signal and Antitumor Effects in Endometrial Cancer through Transcription Factor SP-1.

Long non-coding RNAs (lncRNAs) are implicated in the initiation and progression of a variety of tumors, including endometrial cancer. However, the mechanisms of lncRNA in endometrial cancer formation and progression remain largely unknown. In this study, we confirmed that the lncRNA SNHG4 is upregulated in endometrial cancer and correlates with lower survival rates in endometrial cancer patients. Knock-down of SNHG4 significantly reduced cell proliferation, colonization, migration, and invasion in vitro, as well as modulating the cell cycle and reduced tumor growth of endometrial cancer in vivo. In addition, the effect of SNHG4 by the transcription factor SP-1 was confirmed in vitro. We found in this study that SNHG4/SP-1 plays an important role in endometrial cancer progression and may be used as a potential therapeutic and prognostic biomarker for endometrial cancer.

Tannic acid, an IL-1ß-direct binding compound, ameliorates IL-1ß-induced inflammation and cartilage degradation by hindering IL-1ß-IL-1R1 interaction.

Interleukin-1ß (IL-1ß) is one of the most potent pro-inflammatory cytokines implicated in a wide range of autoinflammatory, autoimmune, infectious, and degenerative diseases. Therefore, many researchers have focused on developing therapeutic molecules that inhibit IL-1ß-IL-1 receptor 1 (IL-1R1) interaction for the treatment of IL-1-related diseases. Among IL-1-related diseases, osteoarthritis (OA), is characterized by progressive cartilage destruction, chondrocyte inflammation, and extracellular matrix (ECM) degradation. Tannic acid (TA) has been proposed to have multiple beneficial effects, including anti-inflammatory, anti-oxidant, and anti-tumor activities. However, it is unclear whether TA plays a role in anti-IL-1ß activity by blocking IL-1ß-IL-1R1 interaction in OA. In this study, we report the anti-IL-1ß activity of TA in the progression of OA in both in vitro human OA chondrocytes and in vivo rat OA models. Herein, using-ELISA-based screening, natural compound candidates capable of inhibiting the IL-1ß-IL-1R1 interaction were identified. Among selected candidates, TA showed hindering IL-1ß-IL-1R1 interaction by direct binding to IL-1ß using surface plasmon resonance (SPR) assay. In addition, TA inhibited IL-1ß bioactivity in HEK-Blue IL-1-dependent reporter cell line. TA also inhibited IL-1ß-induced expression of inducible nitric oxide synthase (NOS2), cyclooxygenase-2 (COX-2), IL-6, tumor necrosis factor-alpha (TNF-α), nitric oxide (NO), and prostaglandin E2 (PGE2) in human OA chondrocytes. Moreover, TA downregulated IL-1ß-stimulated matrix metalloproteinase (MMP)3, MMP13, ADAM metallopeptidase with thrombospondin type 1 motif (ADAMTS)4, and ADAMTS5, while upregulating collagen type II (COL2A1) and aggrecan (ACAN). Mechanistically, we confirmed that TA suppressed IL-1ß-induced MAPK and NF-κB activation. The protective effects of TA were also observed in a monosodium iodoacetamide (MIA)-induced rat OA model by reducing pain and cartilage degradation and inhibiting IL-1ß-mediated inflammation. Collectively, our results provide evidence that TA plays a potential role in OA and IL-1ß-related diseases by hindering IL-1ß-IL-1R1 interaction and suppressing IL-1ß bioactivity.

Butein Inhibits Cell Growth by Blocking the IL-6/IL-6Rα Interaction in Human Ovarian Cancer and by Regulation of the IL-6/STAT3/FoxO3a Pathway.

Butea monosperma (Fabaceae) has been used in traditional Indian medicine to treat a variety of ailments, including abdominal tumors. We aimed to investigate the anti-IL-6 activity of butein in ovarian cancer and elucidate the underlying molecular mechanisms. Butein was isolated and identified from B. monosperma flowers, and the inhibition of IL-6 signaling was investigated using the HEK-Blue™ IL-6 cell line. The surface plasmon resonance assay was used to estimate the binding of butein to IL-6, IL-6Rα, and gp130. After treatment with butein, ovarian cancer cell migration, apoptosis, and tumor growth inhibition were evaluated in vitro and in vivo. Furthermore, we used STAT3 siRNA to identify the mechanistic effects of butein on the IL-6/STAT3/FoxO3a pathway. Butein suppressed downstream signal transduction through higher binding affinity to IL-6. In ovarian cancer, butein inhibited cell proliferation, migration, and invasion, and induced cell cycle arrest and apoptosis. In addition, it decreased the growth of ovarian cancer cells in xenograft tumor models. Butein inhibited STAT3 phosphorylation and induced FoxO3a accumulation in the nucleus by inhibiting IL-6 signaling. The anticancer activity of butein was mediated by blocking the IL-6/IL-6Rα interaction and suppressing IL-6 bioactivity via interfering with the IL-6/STAT3/FoxO3a pathway.

Chelerythrine, a novel small molecule targeting IL-2, inhibits melanoma progression by blocking the interaction between IL-2 and its receptor.

AIMS: In this study, we investigated the inhibition of IL-2 activity and anticancer efficacy of chelerythrine (CHE), a natural small molecule that targets IL-2 and inhibits CD25 binding, and elucidated the mechanism underlying the action of CHE on immune cells. MAIN METHODS: CHE was discovered by competitive binding ELISA and SPR analysis. The effect of CHE on IL-2 activity was evaluated in CTLL-2, HEK-Blue reporter and immune cells, and in ex vivo generation of regulatory T cells (Treg cells). The antitumor activity of CHE was evaluated in B16F10 tumor-bearing C57BL/6 or BALB/c nude mice. KEY FINDINGS: We identified that CHE, which acts as an IL-2 inhibitor, selectively inhibits the interaction between IL-2 and IL-2Rα and directly binds to IL-2. CHE inhibited the proliferation and signaling of CTLL-2 cells and suppressed IL-2 activity in HEK-Blue reporter and immune cells. CHE prevented the conversion of naive CD4+ T cells into CD4+CD25+Foxp3+ Treg cells in response to IL-2. CHE reduced tumor growth in C57BL/6 mice but not in T-cell-deficient mice, upregulated the expression of IFN-γ and cytotoxic molecules, and limited Foxp3 expression. Furthermore, the combination of CHE and a PD-1 inhibitor synergistically increased antitumor activity in melanoma-bearing mice and almost completely regressed the implanted tumors. SIGNIFICANCE: We found that CHE, which targets IL-2 and inhibits its binding to CD25, exhibits T cell-mediated antitumor activity and that combination therapy with CHE and PD-1 inhibitor induced synergistic antitumor effects, suggesting that CHE may be a promising anticancer agent for melanoma monotherapy and combination therapy.

Inhibition of GP130/STAT3 and EMT by combined bazedoxifene and paclitaxel treatment in ovarian cancer.

The interleukin 6 (IL­6)/glycoprotein 130 (GP130)/signal transducer and activator of transcription 3 (STAT3) signalling pathway, with GP130 as an intermediate membrane receptor, is involved in the survival, metastasis, and resistance of ovarian cancer. Bazedoxifene, an FDA­approved drug, is an inhibitor of GP130 and a selective estrogen modulator (SERM). We studied the mechanism of the combination therapy of bazedoxifene and paclitaxel in inhibiting the IL­6­mediated GP130/STAT3 signaling pathway in ovarian cancer. Surface plasmon resonance (SPR) was used to assess the binding of bazedoxifene to GP130. Migration, invasion, and apoptosis of ovarian cancer cells were assessed using bazedoxifene and paclitaxel. In addition, we determined the effects of bazedoxifene and paclitaxel alone or in combination on the GP130/STAT3 pathway and epithelial­mesenchymal transition (EMT). The results revealed that the combination of bazedoxifene and paclitaxel suppressed cell viability, migration, and invasion in the ovarian cancer cells. In addition, the combination treatment increased apoptosis. Furthermore, bazedoxifene combined with paclitaxel inhibited the growth of ovarian cancer cells in a xenograft tumour model. This combination reduced STAT3 phosphorylation and suppressed gene expression and EMT. In conclusion, inhibition of GP130/STAT3 signalling and EMT via a combination of bazedoxifene and paclitaxel could be used as a therapeutic strategy by which to overcome ovarian cancer.

LncRNA SRA mediates cell migration, invasion, and progression of ovarian cancer via NOTCH signaling and epithelial-mesenchymal transition.

Long non-coding RNA (lncRNA) is a newly identified regulator of tumor formation and tumor progression. The function and expression of lncRNAs remain to be fully elucidated, but recent studies have begun to address their importance in human health and disease. The lncRNA, SRA, known as steroid receptor activator, acts as an important modulator of gynecological cancer, and its expression may affect biological functions including proliferation, apoptosis, steroid formation, and muscle development. However, it is still not well known whether SRA is involved in the regulation of ovarian cancer. The present study investigated the molecular function and association between SRA expression and clinicopathological factors. In ovarian cancer cell lines, SRA knockdown and overexpression regulated cell migration, proliferation, and invasion. Both in vivo and in vitro experiments using knockdown and overexpression showed that SRA potently regulated epithelial-mesenchymal transition (EMT) and NOTCH pathway components. Further, clinical data confirmed that SRA was a significant predictor of overall survival (OS) and progression-free survival and patients with ovarian cancer exhibiting high expression of SRA exhibited higher recurrence rates than patients with low SRA expression. In conclusion, the present study indicates that SRA has clinical significance as its expression can predict the prognosis of ovarian cancer patients. High expression of the lncRNA SRA is strongly correlated with recurrence-free survival of ovarian cancer patients.

Bazedoxifene, a GP130 Inhibitor, Modulates EMT Signaling and Exhibits Antitumor Effects in HPV-Positive Cervical Cancer.

Persistent HPV (Human Papillomavirus) infection is the primary cause of cervical cancer. Despite the development of the HPV vaccine to prevent infections, cervical cancer is still a fatal malignant tumor and metastatic disease, and it is often difficult to treat, so a new treatment strategy is needed. The FDA-approved drug Bazedoxifene is a novel inhibitor of protein-protein interactions between IL-6 and GP130. Multiple ligand simultaneous docking and drug repositioning approaches have demonstrated that an IL-6/GP130 inhibitor can act as a selective estrogen modulator. However, the molecular basis for GP130 activation in cervical cancer remains unclear. In this study, we investigated the anticancer properties of Bazedoxifene in HPV-positive cervical cancer cells. In vitro and in vivo experiments showed that Bazedoxifene inhibited cell invasion, migration, colony formation, and tumor growth in cervical cancer cells. We also confirmed that Bazedoxifene inhibits the GP130/STAT3 pathway and suppresses the EMT (Epithelial-mesenchymal transition) sub-signal. Thus, these data not only suggest a molecular mechanism by which the GP130/STAT3 pathway may promote cancer, but also may provide a basis for cervical cancer replacement therapy.

Combination of LMT-28 and Metformin Improves Beneficial Anti-Inflammatory Effect in Collagen-Induced Arthritis.

OBJECTIVES: The interleukin-6 (IL-6)-mediated signaling pathway plays an essential role in the development of rheumatoid arthritis. LMT-28 suppresses the activation of the IL-6-mediated signaling by direct targeting of gp130. Although LMT-28 and metformin both possess anti-inflammatory activity, the beneficial effect of LMT-28 and metformin combination on a collagen-induced arthritis (CIA) model has not yet been investigated. This study aimed to investigate the anti-inflammatory effect and mechanism of a combination of LMT-28 and metformin in a CIA model. METHODS: In MH7A cells, cell proliferation and the IL-6-mediated signaling pathway following administration of LMT-28 and metformin combination was analyzed through MTT assay and Western blotting. The level of T helper 17 (Th17) cell differentiation from CD4+ T cells was analyzed in mouse splenocytes and human peripheral blood mononuclear cells. Arthritis score, incidence rate, inflammatory cytokine, and T-cell subsets were measured in CIA mice following administration of LMT-28 and metformin combination. RESULTS: Combination treatment with LMT-28 and metformin diminished proliferation of MH7A cells and IL-6-mediated gp130, STAT3, and ERK signaling more than in individual treatments. Furthermore, the differentiation of CD4+ T cells into Th17 cells was attenuated more by combination treatment with LMT-28 and metformin than individual treatments. The combination of LMT-28 and metformin ameliorated the arthritic score better than individual treatments. The combination significantly reduced tumor necrosis factor and IL-6 levels in the sera and had an anti-inflammatory effect on the distribution of Treg/Th17 cells in the lymph nodes. CONCLUSION: Combination treatment with LMT-28 and metformin significantly ameliorates arthritic symptoms in CIA by suppressing Th17 differentiation and IL-6 signaling.

Long Noncoding RNA E2F4as Promotes Progression and Predicts Patient Prognosis in Human Ovarian Cancer.

(1) Background: LncRNAs could be a promising biomarker to predict the prognosis of various cancers. The significance of E2F4antisense lncRNA remains unclear in cancer. In this study, we examined the expression level of E2F4as in the serum of ovarian cancer patients and the functional role of E2F4as. (2) Methods: Serum samples were obtained from 108 OC patients and 32 normal patients to measure the expression of E2F4as in the serum. Ovarian cancer cells were used to investigate the role of E2F4as in cell proliferation, invasion, migration and apoptosis, and the expression of E2F4as was knocked down using RNA interference. In addition, E2F4as knockdown cell lines were used in in vivo experiments. (3) Results: The expression of E2F4as was significantly higher in the serum of OC patients than in that of control patients (p < 0.05). The knockdown of E2F4as in ovarian cancer cells led to a decrease in cell proliferation, invasion and migration and an increase in apoptosis. E2F4as knockdown also reduced the expression of epithelium-mesenchymal metastasis (EMT) genes. (4) Conclusion: These findings highlight the clinical significance of E2F4as in predicting the prognosis of OC patients and suggest its potential in promoting tumour aggressiveness by the regulation of EMT-related mechanisms.

Direct Binding to NLRP3 Pyrin Domain as a Novel Strategy to Prevent NLRP3-Driven Inflammation and Gouty Arthritis.

OBJECTIVE: The NLRP3 inflammasome is closely linked to the pathophysiology of a wide range of inflammatory diseases. This study was undertaken to identify small molecules that directly bind to NLRP3 in order to develop pharmacologic interventions for NLRP3-related diseases. METHODS: A structure-based virtual screening analysis was performed with ~62,800 compounds to select efficient NLRP3 inhibitors. The production of caspase 1-p10 and interleukin-1ß (IL-1ß) was measured by immunoblotting and enzyme-linked immunosorbent assay to examine NLRP3 inflammasome activation. Two gouty arthritis models and an air pouch inflammation model induced by monosodium urate monohydrate (MSU) crystal injection were used for in vivo experiments. Primary synovial fluid cells from gout patients were used to determine the relevance of NLRP3 inflammasome inhibition in human gout. RESULTS: Beta-carotene (provitamin A) suppressed the NLRP3 inflammasome activation induced by various activators, including MSU crystals, in mouse bone marrow-derived primary macrophages (P < 0.05). Surface plasmon resonance analysis demonstrated the direct binding of ß-carotene to the pyrin domain (PYD) of NLRP3 (KD = 3.41 × 10-6 ). Molecular modeling and mutation assays revealed the interaction mode between ß-carotene and the NLRP3 PYD. Inflammatory symptoms induced by MSU crystals were attenuated by oral administration of ß-carotene in gouty arthritis mouse models (P < 0.05), correlating with its suppressive effects on the NLRP3 inflammasome in inflamed tissues. Furthermore, ß-carotene reduced IL-1ß secretion from human synovial fluid cells isolated from gout patients (P < 0.05), showing its inhibitory efficacy in human gout. CONCLUSION: Our results present ß-carotene as a selective and direct inhibitor of NLRP3, and the binding of ß-carotene to NLRP3 PYD as a novel pharmacologic strategy to combat NLRP3 inflammasome-driven diseases, including gouty arthritis.

E2F8 regulates the proliferation and invasion through epithelial-mesenchymal transition in cervical cancer.

The transcription factor E2F is an important modulator of the cell cycle, and the unrestricted activation of E2F-dependent transcription is considered to be an important driver of tumor formation and progression. E2F8 is known to play an important role in embryonic development and cell cycle control by inhibiting E2F1. However, it is not yet known whether E2F8 is involved in the progression of cervical cancer. In this study, the functional consequences of E2F8 knockdown in vitro and in vivo were explored. To demonstrate the function of E2F8 in cell proliferation, migration and invasion, we knocked down E2F8 in cervical cancer cell lines; in vitro and in vivo experiments using this knockdown showed that E2F8 potently induced the expression of epithelial-mesenchymal transition (EMT) markers. Finally, clinical data confirmed that E2F8 was a significant predictive factor for progression-free survival, and that patients with cervical cancer who exhibited high expression of E2F8 showed high FIGO stages and frequent recurrence rates compared to patients with low E2F8 expression. In conclusion, our study suggests that E2F8 is highly correlated with the progression-free survival of cervical cancer patients.

Long non-coding RNA steroid receptor activator promotes the progression of endometrial cancer via Wnt/ ß-catenin signaling pathway.

Rationale: Steroid receptor activator (SRA), a long non-coding RNA, serves as a critical regulator of gynecologic cancer. The objective of this study was to determine biological function and clinical significance of SRA expression in endometrial cancer. Method: We investigated whether SRA was involved in the development of endometrial cancer via binding to eukaryotic translation initiation factor 4E-binding protein 1 (EIF4E-BP1) as a transcription factor to enhance Wnt/ ß-catenin signaling pathway. Results: Expression levels of SRA were upregulated in endometrial cancer tissues compared to those in adjacent control tissues. We also found high expression of SRA in EC cells. The relationship between SRA and EIF4E-BP1 was corroborated by transfection of a luciferase reporter plasmid. In addition, SRA knockdown inhibited the expression of EIF4E-BP1 known to play a critical role in the control of protein synthesis, cell growth, and cell survival, thus promoting tumourigenesis and epithelial-mesenchymal transition (EMT) important for cell motility and metastasis. Consistently, immunostaining and western blotting analysis showed that expression levels of ß-catenin and 4EBP1 in the nucleus were significantly decreased by SRA knockdown but increased by SRA over-expression. Conclusions: These results suggest that SRA is involved in proliferation, migration, and invasion of endometrial cancer cells by increasing the expression of EIF4E-BP1 and activity of Wnt/ ß-catenin signaling. These findings indicate that SRA might be a novel biomarker for predicting recurrence and prognosis. It might also serve as a promising therapeutic target in endometrial cancer.

Combination of gp130-targeting and TNF-targeting small molecules in alleviating arthritis through the down-regulation of Th17 differentiation and osteoclastogenesis.

Rheumatoid arthritis (RA) is a systemic, chronic inflammatory disease that is characterized by T helper 17 (Th17) cell- and osteoclast-induced joint destruction and inflammation. In RA, several cytokines (interleukin (IL)-1, 6,17, and tumor necrosis factor (TNF)) are involved in almost all aspects of articular inflammation and destruction. This study aimed to evaluate the combinatorial effect of TNF and IL-6 inhibitors on the differentiation and activation of Th17 cells and osteoclasts in the context of RA, and to identify the RA-related mechanisms through IL-6 signaling. Tetrahydropapaverine (THP) showed direct binding to TNF in screening-ELISA, and SPR and TNF-neutralization assays. In a previous study, the therapeutic effect of gp130-targeting LMT-28 was confirmed in RA. Combinatorial treatment with LMT-28 and THP reduced the arthritis index and showed protective effects against bone and cartilage destruction in CIA mice. The secretion levels of TNF, IL-6, and IL-1ß significantly decreased upon combinatorial treatment with LMT-28 and THP. Further, the LMT-28 and THP combination suppressed the differentiation and activation of Th17 cells in mouse splenocytes and human PBMCs. In human RA-FLS, the LMT-28 and THP combination inhibited cell proliferation and downregulated IL-6 and/or TNF-mediated signaling relative to that observed upon independent treatment with LMT-28 or THP. Furthermore, the combination of LMT-28 and THP significantly inhibited the differentiation of mouse bone marrow monocytes (BMMs) into osteoclasts. In conclusion, the LMT-28 and THP combination can attenuate RA through the inhibition of Th17 differentiation and osteoclastogenesis, and suppression of IL-6 or TNF-induced signaling pathways. This combinatorial therapy could be used as a new strategy for the treatment of RA.