Clinical significance of ZHX2 in colitis and colorectal carcinogenesis.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. e15010-e15010
Author(s):  
Yonghua Bao ◽  
Yongchen Guo ◽  
Wancai Yang
Keyword(s):  
Mouse Model ◽  
Malignant Transformation ◽  
Clinical Significance ◽  
Proteomic Analysis ◽  
Colorectal Carcinogenesis ◽  
Beta Catenin ◽  
Mrna Levels ◽  
Chronic Colitis ◽  
Rna Sequence

e15010 Background: Colorectal cancer (CRC) is one of the most common malignancies. Mechanistic studies have revealed that the malignant transformation of chronic colitis is one of the major causes besides the activation of Wnt/beta-catenin signaling. Our preliminary data have shown that ZHX2 is a common target of the miRNAs and is associated with colitis malignant transformation. This study aimed to explore the changes of ZHX2 in colitis-CRC tissues and their clinical significance, and to reveal the underlying mechanism. Methods: ZHX2 protein expression levels were analyzed by immunohistochemical staining in a CRC tissue microarray containing 400 cases of CRC and matched normal mucosa; mRNA levels were analyzed in fresh tissues by qRT-PCR. ZHX2 functional studies were conducted in ZHX2 transgenic mouse model and in vitro; Protein regulation was assayed by proteomic analysis using iTRAQ and RNA changes were assayed by RNA-sequence. Results: Protein and mRNA levels of ZHX2 were significantly increased in CRC and was associated with poor outcomes and recurrence, the copy numbers were also elevated in CRC. Intestine-specific transgenic Zhx2 mouse model showed chronic inflammation in the intestine at early age and intestinal carcinogenesis at late ages. Intestinal epithelial cell RNA sequence assay showed differential expression of genes and signaling pathways, compared to the wild-type mice. Biological functional studied showed that increasing ZHX2 in CRC cells enhanced cell proliferation, motility and migration, and suppressed cell apoptosis. Proteomic analysis showed that ZHX2 and HMGA1 were synergistic during tumorigenesis; mi-RNA regulatory study showed that miR-150 targeted ZHX2 and suppressed ZHX2 expression, but ZHX2 could regulate Wnt/beta-catenin and Serine family member PRSS8. Conclusions: This study has revealed the critical roles of ZHX2 in chronic colitis and its malignant transformation and underlying molecular mechanism, identifying a novel target for CRC prevention and therapy.

scholarly journals PRKAR1B-AS2 Long Noncoding RNA Promotes Tumorigenesis, Survival, and Chemoresistance via the PI3K/AKT/mTOR Pathway

2021 ◽  
Vol 22 (4) ◽  
pp. 1882
Author(s):  
Abdelrahman M. Elsayed ◽  
Emine Bayraktar ◽  
Paola Amero ◽  
Salama A. Salama ◽  
Abdelaziz H. Abdelaziz ◽  
...  
Keyword(s):  
Mouse Model ◽  
Long Noncoding Rna ◽  
Proteomic Analysis ◽  
Noncoding Rna ◽  
Mtor Pathway ◽  
The Cancer Genome Atlas ◽  
Migration And Invasion ◽  
Physical Interaction ◽  
Cisplatin Sensitivity

Many long noncoding RNAs have been implicated in tumorigenesis and chemoresistance; however, the underlying mechanisms are not well understood. We investigated the role of PRKAR1B-AS2 long noncoding RNA in ovarian cancer (OC) and chemoresistance and identified potential downstream molecular circuitry underlying its action. Analysis of The Cancer Genome Atlas OC dataset, in vitro experiments, proteomic analysis, and a xenograft OC mouse model were implemented. Our findings indicated that overexpression of PRKAR1B-AS2 is negatively correlated with overall survival in OC patients. Furthermore, PRKAR1B-AS2 knockdown-attenuated proliferation, migration, and invasion of OC cells and ameliorated cisplatin and alpelisib resistance in vitro. In proteomic analysis, silencing PRKAR1B-AS2 markedly inhibited protein expression of PI3K-110α and abrogated the phosphorylation of PDK1, AKT, and mTOR, with no significant effect on PTEN. The RNA immunoprecipitation detected a physical interaction between PRKAR1B-AS2 and PI3K-110α. Moreover, PRKAR1B-AS2 knockdown by systemic administration of 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine nanoparticles loaded with PRKAR1B-AS2–specific small interfering RNA enhanced cisplatin sensitivity in a xenograft OC mouse model. In conclusion, PRKAR1B-AS2 promotes tumor growth and confers chemoresistance by modulating the PI3K/AKT/mTOR pathway. Thus, targeting PRKAR1B-AS2 may represent a novel therapeutic approach for the treatment of OC patients.

scholarly journals Daphnetin inhibits proliferation and inflammatory response in human HaCaT keratinocytes and ameliorates imiquimod-induced psoriasis-like skin lesion in mice

2020 ◽  
Vol 53 (1) ◽  
Author(s):  
Jintao Gao ◽  
Fangru Chen ◽  
Huanan Fang ◽  
Jing Mi ◽  
Qi Qi ◽  
...  
Keyword(s):  
Skin Lesion ◽  
Inflammatory Response ◽  
Mouse Model ◽  
Nuclear Translocation ◽  
Marker Gene ◽  
Inflammatory Factors ◽  
Mrna Levels ◽  
Hacat Keratinocytes ◽  
Tnf Α

Abstract Background Psoriasis is a common chronic inflammatory skin disease. Keratinocytes hyperproliferation and excessive inflammatory response contribute to psoriasis pathogenesis. The agents able to attenuate keratinocytes hyperproliferation and excessive inflammatory response are considered to be potentially useful for psoriasis treatment. Daphnetin exhibits broad bioactivities including anti-proliferation and anti-inflammatory. This study aims to evaluate the anti-psoriatic potential of daphnetin in vitro and in vivo, and explore underlying mechanisms. Methods HaCaT keratinocytes was stimulated with the mixture of IL-17A, IL-22, oncostatin M, IL-1α, and TNF-α (M5) to establish psoriatic keratinocyte model in vitro. Cell viability was measured using Cell Counting Kit-8 (CCK-8). Quantitative Real-Time PCR (qRT-PCR) was performed to measure the mRNA levels of hyperproliferative marker gene keratin 6 (KRT6), differentiation marker gene keratin 1 (KRT1) and inflammatory factors IL-1β, IL-6, IL-8, TNF-α, IL-23A and MCP-1. Western blotting was used to detect the protein levels of p65 and p-p65. Indirect immunofluorescence assay (IFA) was carried out to detect p65 nuclear translocation. Imiquimod (IMQ) was used to construct psoriasis-like mouse model. Psoriasis severity (erythema, scaling) was scored based on Psoriasis Area Severity Index (PASI). Hematoxylin and eosin (H&E) staining was performed to examine histological change in skin lesion. The expression of inflammatory factors including IL-6, TNF-α, IL-23A and IL-17A in skin lesion was measured by qRT-PCR. Results Daphnetin attenuated M5-induced hyperproliferation in HaCaT keratinocytes. M5 stimulation significantly upregulated mRNA levels of IL-1β, IL-6, IL-8, TNF-α, IL-23A and MCP-1. However, daphnetin treatment partially attenuated the upregulation of those inflammatory cytokines. Daphnetin was found to be able to inhibit p65 phosphorylation and nuclear translocation in HaCaT keratinocytes. In addition, daphnetin significantly ameliorate the severity of skin lesion (erythema, scaling and epidermal thickness, inflammatory cell infiltration) in IMQ-induced psoriasis-like mouse model. Daphnetin treatment attenuated IMQ-induced upregulation of inflammatory cytokines including IL-6, IL-23A and IL-17A in skin lesion of mice. Conclusions Daphnetin was able to attenuate proliferation and inflammatory response induced by M5 in HaCaT keratinocytes through suppression of NF-κB signaling pathway. Daphnetin could ameliorate the severity of skin lesion and improve inflammation status in IMQ-induced psoriasis-like mouse model. Daphnetin could be an attractive candidate for future development as an anti-psoriatic agent.

scholarly journals A Novel Low-Risk Germline Variant in the SH2 Domain of the SRC Gene Affects Multiple Pathways in Familial Colorectal Cancer

2021 ◽  
Author(s):  
Diamanto Skopelitou ◽  
Beiping Miao ◽  
Aayushi Srivastava ◽  
Abhishek Kumar ◽  
Magdalena Kuświk ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cancer Susceptibility ◽  
In Silico Analysis ◽  
Missense Variant ◽  
Sh2 Domain ◽  
Colorectal Carcinogenesis ◽  
Beta Catenin ◽  
Mrna Levels ◽  
Potential Marker ◽  
Src Gene

Colorectal cancer (CRC) shows one of the largest proportions of familial cases among different malignancies, but only 5-10% of all CRC cases are linked to mutations in established predisposition genes. Thus, familial CRC constitutes a promising target for the identification of novel, high- to moderate-penetrance germline variants underlying cancer susceptibility by next generation sequencing. In this study, we performed whole genome sequencing on 3 members of a family with CRC aggregation. Subsequent integrative in silico analysis using our in-house developed variant prioritization pipeline resulted in the identification of a novel germline missense variant in SRC gene (V177M), a proto-oncogene highly upregulated in CRC. Functional validation experiments in HT-29 cells showed that introduction of SRCV177M resulted in increased cell proliferation and enhanced protein expression of phospho-SRC (Y419), a potential marker for SRC activity. Upregulation of paxillin, β-Catenin and STAT3 mRNA levels, increased levels of phospho-ERK, CREB and CCND1 proteins and downregulation of the tumor suppressor p53 further proposed the activation of several pathways due to the SRCV177M variant. The findings of our pedigree-based study contribute to the exploration of the genetic background of familial CRC and bring insights into the molecular basis of upregulated SRC activity and downstream pathways in colorectal carcinogenesis.

scholarly journals Therapeutic Targeting of Nrf2 Signaling by Maggot Extracts Ameliorates Inflammation-Associated Intestinal Fibrosis in Chronic DSS-Induced Colitis

2021 ◽  
Vol 12 ◽  
Author(s):  
Rong Wang ◽  
Daojuan Wang ◽  
Hongwei Wang ◽  
Tingyu Wang ◽  
Yajing Weng ◽  
...  
Keyword(s):  
Pathological Feature ◽  
Mrna Levels ◽  
Protective Effects ◽  
Clinical Markers ◽  
Chronic Colitis ◽  
Intestinal Fibrosis ◽  
Nrf2 Expression ◽  
Tgf Β1

Intestinal fibrosis is induced by excessive myofibroblast proliferation and collagen deposition, which has been regarded as a general pathological feature in inflammatory bowel disease (IBD). Therefore, identifying clinical markers and targets to treat and prevent intestinal fibrosis is urgently needed. The traditional Chinese medicine maggot, commonly known as “wu gu chong”, has been shown to reduce oxidative stress and alleviate inflammation in chronic colitis. This study investigated the mechanisms underlying the effects of maggot extract (ME) on inflammation-associated intestinal fibrosis in TGF-β1-stimulated human intestinal fibroblasts (CCD-18Co cells) and dextran sodium sulphate (DSS)-induced chronic colitis murine model. To assess the severity of inflammation and fibrosis, histological and macroscopic evaluation were carried out. The results showed that ME was a significant inhibitor of body weight loss and colon length shortening in mice with chronic colitis. In addition, ME suppressed the intestinal fibrosis by downregulating TGF-β1/SMADs pathway via upregulation of Nrf2 expression at both protein and mRNA levels. ME markedly increased the expression of Nrf2, thus resulting in a higher level of HO-1. After treatment with Nrf2 inhibitor (ML385) or siRNA-Nrf2 for deactivating Nrf2 pathway, the protective effects of ME were abolished both in vitro and in vivo. Moreover, the histopathological results for the major organs of DSS mice treated with ME showed no signs of clinically important abnormalities. Treatment with ME had no effect on the viability of CCD-18Co cells, suggesting its low in vitro cytotoxicity. Furthermore, ME could mediate intestine health by keeping the balance of the gut microbes through the enhancement of beneficial microbes and suppression of pathogenic microbes. In conclusion, this is the first ever report demonstrating that ME ameliorates inflammation-associated intestinal fibrosis by suppressing TGF-β1/SMAD pathway via upregulation of Nrf2 expression. Our findings highlight the potential of Nrf2 as an effective therapeutic target for alleviating intestinal fibrosis.

scholarly journals MicroRNA-194: a novel regulator of glucagon-like peptide-1 synthesis in intestinal L cells

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jiao Wang ◽  
Di Zhao ◽  
Cheng-Zhi Ding ◽  
Feng Guo ◽  
Li-Na Wu ◽  
...  
Keyword(s):  
Mrna Level ◽  
Beta Catenin ◽  
Mrna Levels ◽  
Obese Mice ◽  
L Cells ◽  
The Status ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1 ◽  
Induced Apoptosis

AbstractIn the status of obesity, the glucagon-like peptide-1 (GLP-1) level usually declines and results in metabolic syndrome. This study aimed to investigate the intracellular mechanism of GLP-1 synthesis in L cells from the perspective of microRNA (miRNA). In the present study, we found that GLP-1 level was down-regulated in the plasma and ileum tissues of obese mice, while the ileac miR-194 expression was up-regulated. In vitro experiments indicated that miR-194 overexpression down-regulated GLP-1 level, mRNA levels of proglucagon gene (gcg) and prohormone convertase 1/3 gene (pcsk1), and the nuclear protein level of beta-catenin (β-catenin). Further investigation confirmed that β-catenin could promote gcg transcription through binding to transcription factor 7-like 2 (TCF7L2). miR-194 suppressed gcg mRNA level via negatively regulating TCF7L2 expression. What’s more, forkhead box a1 (Foxa1) could bind to the promoter of pcsk1 and enhanced its transcription. miR-194 suppressed pcsk1 transcription through targeting Foxa1. Besides, the interference of miR-194 reduced palmitate (PA)-induced cell apoptosis and the anti-apoptosis effect of miR-194 inhibitor was abolished by TCF7L2 knockdown. Finally, in HFD-induced obese mice, the silence of miR-194 significantly elevated GLP-1 level and improved the metabolic symptoms caused by GLP-1 deficiency. To sum up, our study found that miR-194 suppressed GLP-1 synthesis in L cells via inhibiting TCF7L2-mediated gcg transcription and Foxa1-mediated pcsk1 transcription. Meanwhile, miR-194 took part in the PA-induced apoptosis of L cells.

scholarly journals The Cold-Shock Protein Ybx1 Is Required for Development and Maintenance of Acute Myeloid Leukemia (AML) in Vitro and In Vivo

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 792-792
Author(s):  
Florian Perner ◽  
Ashok K. Jayavelu ◽  
Tina Maria Schnoeder ◽  
Nomusa Mashamba ◽  
Juliane Mohr ◽  
...  
Keyword(s):  
Mouse Model ◽  
Protein Expression ◽  
Malignant Transformation ◽  
Cell Lines ◽  
Median Survival ◽  
Cold Shock ◽  
Rna Binding ◽  
Cold Shock Protein ◽  
Significant Delay

Abstract The family of cold shock proteins (CSPs) is highly conserved and consists of 8 members, including Ybx1-3, Csde1 and Lin28. Ybx1 is a multifunctional DNA/RNA binding protein that modulates gene transcription and translation during inflammation and malignant transformation. Recently, our group identified Ybx1 as a mediator of Jak2 signaling in MPN that protects Jak2-mutated cells from Jak-inhibitor induced apoptosis. In a recently published genome wide CRISPR-Cas9 dropout screen in AML cell lines, depletion of Ybx1 resulted in the highest dropout indices compared to other CSP members, with strongest dependencies in cell lines harboring MLL-rearrangements. Protein expression of Ybx1 in healthy individuals (n=10), primary MDS (n=54) and AML (n=58) bone marrow (BM) biopsies, revealed high protein expression in the majority of AML and MDS cases. Consistently, gene expression data revealed high mRNA expression of Ybx1 in AML samples compared to normal controls. Genetic inactivation of Ybx1 in human AML cell lines by RNAi resulted in reduced proliferative capacity. Therefore, we sought to investigate the requirement for Ybx1 in malignant transformation. We used BM cells from a previously published conventional knockout (ko) mouse model (Lu et al., 2005) in which homozygous deletion is embryonically lethal due to brain malformation. We sorted Lineage-Sca1+Kit+ (LSK-) cells from the BM of heterozygous (Ybx1+/-) and wildtype (Ybx1+/+) mice. Cells were retrovirally infected with either MLL-AF9 (MA9) or HoxA9 and Meis1a (HA9M1) to assess for disease development by serial plating in methylcellulose. Haploinsufficiency for Ybx1 in MA9- or HA9M1 transformed cells limited re-plating capacity to 2-4 rounds. When we injected 2,5x 104 MA9-infected LSK cells into sublethally irradiated recipient mice, recipients of MA9-Ybx1+/- cells (n=8) and MA9-Ybx1+/+ (n=10) showed development of AML. However, recipients of MA9-Ybx1+/- cells had a significant delay in AML development (median survival 67.5 days for Ybx1+/+ versus 101.5 days for Ybx1+/- animals, p=0.0078**). This effect appeared even more pronounced when 1x 106 whole BM cells were transplanted into sublethally irradiated secondary recipients. Besides a significant delay in AML development (median survival 37.5 days for recipients of MA9-Ybx1+/+ versus 79 days for MA9-Ybx1+/- BM, p=0.0042**), disease penetrance was reduced by 40%, indicating that haploinsufficiency for Ybx1 impairs development of MA9 driven AML. In contrast, immunophenotypic abundance of stem- and progenitor cells in Ybx1+/+ versus Ybx1+/- animals revealed comparable numbers in all relevant subpopulations. Serial competitive transplantation of Ybx1+/+ and Ybx1+/- BM into primary and secondary recipient animals showed no competitive disadvantage or lack of self-renewal capacity of Ybx1+/- cells. To address the question whether Ybx1 may also be essential for maintenance of AML, we used RNAi to deplete Ybx1 in already established MA9 driven AML. LSK cells from BL/6 mice transformed with MA9 were injected into primary recipient mice. After AML onset, MA9-LSK cells were sorted and infected with either one of 3 shRNAs against Ybx1 or non-targeting (NT-) control. Lentiviral knockdown of 40% reduced colony formation by more than 50% but did not limit the re-plating capacity in vitro. When injected into sub-lethally irradiated recipient mice, lentiviral knockdown (kd) of Ybx1 resulted in a significant delay in AML development (median survival 39.5 days for NT-control versus 53 days for Ybx1 kd, p=0.0446*). To validate our findings, we used a newly generated conditional ko mouse model for Ybx1, in which exon 3 coding for the cold-shock domain is deleted by activation of an Mx1-Cre-recombinase following pIpC administration. Preliminary results provide first evidence that genetic deletion of Ybx1 after onset of MA9 driven leukemia resulted in improved survival of primary recipient (median survival 73 versus 83 days) and a reduced penetrance in secondary recipient mice. Taken together our results may provide first evidence for a functional role of Ybx1 in MLL-AF9 driven AML. As Ybx1 seems to be dispensable for normal hematopoietic cells, these findings may offer a potential therapeutic index. Experiments to assess for the requirement for Ybx1 in maintenance of murine and human AML as well as analysis on proteomic and transcriptional changes following Ybx1 deletion are currently under way. Disclosures No relevant conflicts of interest to declare.

scholarly journals Inhibitory effect of selenomethionine on carcinogenesis in the model of human colorectal cancer in vitro and its link to the Wnt/β-catenin pathway.

2018 ◽  
Vol 65 (3) ◽  
Author(s):  
Edyta Korbut ◽  
Agata Ptak-Belowska ◽  
Tomasz Brzozowski
Keyword(s):  
Colorectal Cancer ◽  
Cancer Cells ◽  
Colorectal Carcinogenesis ◽  
Mrna Levels ◽  
Human Colorectal Cancer ◽  
Bax Protein ◽  
Colorectal Cancer Cells ◽  
Gsk 3Β ◽  
Ht 29

Selenium compounds have been implicated as anticancer agents; however, the mechanism of their inhibitory action against cancer development has not been extensively investigated. The constitutive activation of the Wnt/β-catenin pathway is a central event in colorectal carcinogenesis. In this pathway, the excessive cell proliferation is initiated by the generation of β-catenin followed by overexpression of proto-oncogenes such as c-Myc. It is believed that under physiological conditions the level of c-Myc is efficiently controlled by accessibility of β-catenin protein through the process of phosphorylation by glycogen synthase kinase 3β (GSK-3β). Here, we determined whether selenomethionine (SeMet) can inhibit cell growth and affect the Wnt/β-catenin pathway in HT-29 human colorectal cancer cells in vitro. The effective cytotoxic doses of SeMet have been selected after 48 h of incubation of this compound with colorectal cancer HT-29 cell line. The MTT assay was used to assess cell viability and the protein and mRNA levels of β-catenin and c-Myc were determined by Western blotting and qPCR, respectively. The SeMet potently inhibited growth of HT-29 cells, significantly decreased the β-catenin protein and mRNA concentration, down-regulated the c-Myc gene expression and up-regulated pro-apoptotic Bax protein expression. Moreover, SeMet increased the level of GSK-3β phosphorylated at serine 9 (S9) and significantly increased the level of β-catenin phosphorylated at S33 and S37. We conclude that SeMet suppresses the growth of HT-29 colorectal cancer cells by the mechanism linked to the Wnt/β-catenin pathway, however, the degradation of β-catenin may occur independently of GSK-3β catalytic activity and its phosphorylation status.

scholarly journals In vitro metabolic zonation through oxygen gradient on a chip

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Federica Tonon ◽  
Giovanni Giuseppe Giobbe ◽  
Alessandro Zambon ◽  
Camilla Luni ◽  
Onelia Gagliano ◽  
...  
Keyword(s):  
Embryonic Stem ◽  
Glycogen Storage ◽  
Beta Catenin ◽  
Mrna Levels ◽  
Carbamoyl Phosphate ◽  
Glycogen Accumulation ◽  
Oxygen Gradient ◽  
Metabolic Zonation

Abstract Among the multiple metabolic signals involved in the establishment of the hepatic zonation, oxygen could play a key role. Indeed, depending on hepatocyte position in the hepatic lobule, gene expression and metabolism are differently affected by the oxygen gradient present across the lobule. The aim of this study is to understand whether an oxygen gradient, generated in vitro in our developed device, is sufficient to instruct a functional metabolic zonation during the differentiation of human embryonic stem cells (hESCs) from endoderm toward terminally differentiated hepatocytes, thus mimicking the in vivo situation. For this purpose, a microfluidic device was designed for the generation of a stable oxygen gradient. The oxygen gradient was applied to differentiating hESCs at the pre-hepatoblast stage. The definitive endoderm and hepatic endoderm cells were characterized by the expression of the transcription factor SOX-17 and alpha-fetoprotein (AFP). Immature and mature hepatocytes were characterized by hepatocyte nuclear factor 4-alpha (HNF-4α) and albumin (ALB) expression and also analyzed for cytochrome P450 (CYP3A4) zonation and glycogen accumulation through PAS staining. Metabolic zonated genes expression was assessed through quantitative real time PCR. Application of the oxygen gradient during differentiation induced zonated glycogen storage, which was higher in the hepatocytes grown in high pO2 compared to those grown in low pO2. The mRNA levels of glutamine synthetase (GLUL), beta-catenin (CTNNB) and its direct target cyclin D1 (CCND1) showed significantly higher expression in the cells grown in low pO2 compared to those grown in high pO2. On the contrary, carbamoyl-phosphate synthetase 1 (CPS1), ALB, the proliferative marker ki67 (MKI67) and cyclin A (CCNA) resulted to be significantly higher expressed in cells cultured in high pO2 compared to those cultured in low pO2. These results indicate that the oxygen gradient generated in our device can instruct the establishment of a functional metabolic zonation in differentiating hESCs. The possibility to obtain differentiated hepatocytes in vitro may allow in the future to deepen our knowledge about the physiology/pathology of hepatocytes in relation to the oxygen content.

scholarly journals Benefit of combining curcumin, harpagophytum and bromelain to reduce inflammation in osteoarthritic synovial cells

2021 ◽  
Author(s):  
Sybille Brochard ◽  
Julien Pontin ◽  
Benoit Bernay ◽  
Karim Boumediene ◽  
Thierry Conrozier ◽  
...  
Keyword(s):  
Proteomic Analysis ◽  
Complex Disease ◽  
Inflammatory Diseases ◽  
Biological Action ◽  
Inflammatory Factors ◽  
Synovial Cells ◽  
Mrna Levels ◽  
Expression Of Genes ◽  
Synovial Tissues

Abstract Background: Osteoarthritis is the most common cause of arthritis affecting millions of people worldwide, characterized by joint pain and inflammation. It is a complex disease involving inflammatory factors and affecting the whole joint including synovium. Since drug combination is widely used to treat chronic inflammatory diseases, a similar strategy may be worth of interest to design plant-derived natural products to reduce inflammation in OA joint. Here, we characterized the response of OA synovial cells to lipopolysaccharide (LPS) and investigated the biological action of the combination of curcumin, harpagophytum and bromelain in this original in vitro model of osteoarthritis.Methods: Primary, human synovial cells from OA patients were stimulated with LPS and proteomic analysis was performed. Bioinformatics analysis were performed using Cytoscape App and SkeletalVis databases. Additionally, cells were treated with curcumin, harpagophytum and bromelain alone or the three vegetal compounds together. The expression of genes involved in inflammation, pain or catabolism were determined by RT-PCR. The release of the encoded proteins by these genes and of prostaglandin E2 (PGE2) were also assayed by ELISA. Results: Proteomic analysis demonstrated that LPS induces the expression of numerous proteins involved in OA process in human OA synovial cells. In particular, it stimulates inflammation through the production of pro-inflammatory cytokines (Interleukin-6, IL-6), the catabolism through an increase of metalloproteases (MMP-1, MMP-3, MMP-13), and the production of pain-mediating neurotrophin (Nerve Growth Factor, NGF). These increases were observed at level of mRNA levels and of protein release. LPS also increases the amount of PGE2, another inflammation and pain mediator. At doses tested, vegetal extracts had little effects: only curcumin slightly counteracted the effects of LPS on NGF and MMP13 mRNA, and PGE2, IL-6 and MMP13 release. In contrast the association of curcumin with harpagophytum and bromelain reversed lots of effects of LPS in human OA synovial cells. It significantly reduced the gene expression and/or the release of proteins involved in catabolism (MMP3 and 13), inflammation (IL-6) and pain (PGE2 and NGF).Conclusion: We show that the stimulation of human OA synovial cells with LPS permit to induce protein changes similar to an inflamed OA synovial tissues. In addition, using this model, we demonstrate that the combination of three vegetal compounds, namely curcumin, harpagophytum and bromelain have anti-inflammatory and anti-catabolic action in synovial cells and may thus reduce OA progression and related-pain.

scholarly journals The benefit of combining curcumin, bromelain and harpagophytum to reduce inflammation in osteoarthritic synovial cells

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Sybille Brochard ◽  
Julien Pontin ◽  
Benoit Bernay ◽  
Karim Boumediene ◽  
Thierry Conrozier ◽  
...  
Keyword(s):  
Gene Expression ◽  
Proteomic Analysis ◽  
Complex Disease ◽  
Inflammatory Diseases ◽  
Biological Action ◽  
Inflammatory Factors ◽  
Synovial Cells ◽  
Mrna Levels ◽  
Bioinformatics Analyses

Abstract Background Osteoarthritis (OA) is the most common form of arthritis, affecting millions of people worldwide and characterised by joint pain and inflammation. It is a complex disease involving inflammatory factors and affecting the whole joint, including the synovial membrane. Since drug combination is widely used to treat chronic inflammatory diseases, a similar strategy of designing plant-derived natural products to reduce inflammation in OA joints may be of interest. In this study, we characterised the response of OA synovial cells to lipopolysaccharide (LPS) and investigated the biological action of the combination of curcumin, bromelain and harpagophytum in this original in vitro model of osteoarthritis. Methods Firstly, human synovial cells from OA patients were stimulated with LPS and proteomic analysis was performed. Bioinformatics analyses were performed using Cytoscape App and SkeletalVis databases. Additionally, cells were treated with curcumin, bromelain and harpagophytum alone or with the three vegetal compounds together. The gene expression involved in inflammation, pain or catabolism was determined by RT-PCR. The release of the encoded proteins by these genes and of prostaglandin E2 (PGE2) were also assayed by ELISA. Results Proteomic analysis demonstrated that LPS induces the expression of numerous proteins involved in the OA process in human OA synovial cells. In particular, it stimulates inflammation through the production of pro-inflammatory cytokines (Interleukin-6, IL-6), catabolism through an increase of metalloproteases (MMP-1, MMP-3, MMP-13), and the production of pain-mediating neurotrophins (Nerve Growth Factor, NGF). These increases were observed in terms of mRNA levels and protein release. LPS also increases the amount of PGE2, another inflammation and pain mediator. At the doses tested, vegetal extracts had little effect: only curcumin slightly counteracted the effects of LPS on NGF and MMP-13 mRNA, and PGE2, IL-6 and MMP-13 release. In contrast, the combination of curcumin with bromelain and harpagophytum reversed lots of effects of LPS in human OA synovial cells. It significantly reduced the gene expression and/or the release of proteins involved in catabolism (MMP-3 and -13), inflammation (IL-6) and pain (PGE2 and NGF). Conclusion We have shown that the stimulation of human OA synovial cells with LPS can induce protein changes similar to inflamed OA synovial tissues. In addition, using this model, we demonstrated that the combination of three vegetal compounds, namely curcumin, bromelain and harpagophytum, have anti-inflammatory and anti-catabolic effects in synovial cells and may thus reduce OA progression and related pain.

Sign in / Sign up

Export Citation Format

Share Document