Lactobacillus casei enhances type II collagen/glucosamine-mediated suppression of inflammatory responses in experimental osteoarthritis

AimsWe previously reported that Lactobacillus casei (L. casei) has beneficial effects in experimental rheumatoid arthritis (RA) by suppressing inflammatory immune responses. The major purpose of this study was to evaluate therapeutic effects of L. casei on pathological responses in experimental rodent model of osteoarthritis (OA).Main methodsExperimental OA was induced by intra-articular injection of monosodium iodoacetate (MIA) in Wistar rats. L. casei alone or together with type II collagen (CII) and glucosamine (Gln) was orally administered into OA rats. The pathophysiological aspects of OA were investigated by analyzing mechanical hyperalgesia and histology of articular tissues. Expression of inflammatory molecules was analyzed in CD4⁺ T cells, synovial fibroblasts, and chondrocytes by quantitative real-time PCR.Key findingsOral administration of L. casei together with CII and Gln more effectively reduced pain, cartilage destruction, and lymphocyte infiltration than the treatment of Gln or L. casei alone. This co-administration also decreased expression of various pro-inflammatory cytokines (interleukin-1β (IL-1β), IL-2, IL-6, IL-12, IL-17, tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ)) and matrix metalloproteinases (MMP1, MMP3, and MMP13), while up-regulating anti-inflammatory cytokines (IL-4 and IL-10). These results are concomitant with reduced translocation of NF-κB into the nucleus and increased expression of the tissue inhibitor of MMP1 (TIMP1) and CII in chondrocytes.SignificanceOur study provides evidence that L. casei could act as a potent nutraceutical modulator for OA treatment by reducing pain, inflammatory responses, and articular cartilage degradation.     

Anti-osteoclastic effect of caffeic acid phenethyl ester in murine macrophages depends upon the suppression of superoxide anion production through the prevention of an active-Nox1 complex formation

This study investigated the anti-osteoclastic effect of caffeic acid phenethyl ester (CAPE) through suppression of Nox1-mediated superoxide anions production. The multi-nucleated cells were counted and followed by measuring their tartrate-resistant acid phosphatase (TRAP) activity. The superoxide anion production was determined by using fluorescent probe dihydroethidium (DHE). After one day of exposure to the receptor activator of nuclear factor-κB ligand (RANKL), the expression of the proteins involved in superoxide anion production was determined by western blotting. A potent anti-osteoclastic effect of CAPE was observed; the superoxide anion level reached a maximum value after one day of incubation. CAPE attenuated the expression of NADPH (nicotinamide adenine dinucleotide phosphate) oxidase 1 (Nox1) and Rac1, and mitigated the RANKL-induced translocation of p47^phox to the cell membrane. In addition, CAPE suppressed the expression of nuclear factor-kappa B (NF-κB p65), its translocation to the nucleus, and the activation of NF-κB inhibitor (IκBα) and its kinase (IKKβ). Furthermore, CAPE diminished the expression and activation of the c-jun N-terminal kinase (JNK) and the expression of protein-1 activators (AP-1) such as c-Fos and c-Jun. The expression of Nox1 was suppressed by CAPE through the down-regulation of IKKβ/IκBα/NF-κB and JNK/AP-1 signal pathway. This study provides evidence that the anti-osteoclastic effect of CAPE depends upon the attenuated superoxide anion production, which is closely related with interruption of an active Nox1 complex formation due to the attenuated catalytic subunit Nox1 expression resulting from suppression of the IKKβ/IκBα/NF-κB and JNK/AP-1 signaling pathway and the down-regulation of the p47^phox subunit translocation to the cell membrane.     

Photobiomodulation therapy in knee osteoarthritis reduces oxidative stress and inflammatory cytokines in rats

Knee osteoarthritis (OA) is a chronic disease that causes pain and gradual degeneration of the articular cartilage. In this study, MIA‐induced OA knee model was used in rats to test the effects of the photobiomodulation therapy (PBM). We analyzed the inflammatory process (pain and cytokine levels), and its influence on the oxidative stress and antioxidant capacity. Knee OA was induced by monosodium iodoacetate (MIA) intra‐articular injection (1.5 mg/50 μL) and the rats were treated with eight sessions of PBM 3 days/week (904 nm, 6 or 18 J/cm²). For each animal, mechanical and cold hyperalgesia and spontaneous pain were evaluated; biological analyses were performed in blood serum, intra‐articular lavage, knee structures, spinal cord and brainstem. Cytokine assays were performed in knee, spinal cord and brainstem samples. The effects of the 18 J/cm² dose of PBM were promising in reducing pain and neutrophil activity in knee samples, together with reducing oxidative stress damage in blood serum and spinal cord samples. PBM improved the antioxidant capacity in blood serum and brainstem, and decreased the knee pro‐inflammatory cytokine levels. Our study demonstrated that PBM decreased oxidative damage, inflammation and pain. Therefore, this therapy could be an important tool in the treatment of knee OA.     

The antioxidant edaravone attenuates ER-stress-mediated cardiac apoptosis and dysfunction in rats with autoimmune myocarditis

Abstract

Experimental autoimmune myocarditis (EAM) is mediated by myocardial infiltration by myosin-specific T-cells secreting inflammatory cytokines. In this study, rat models of EAM were prepared by injection with porcine cardiac myosin. One week after immunization, edaravone was administered intraperitoneally at 3 or 10 mg/kg/day to rats for 2 weeks. Cardiac function was measured by haemodynamic and echocardiographic studies and TUNEL assay was performed. Left ventricular (LV) expression of NADPH oxidase sub-units (p47^phox and p67^phox), pro-inflammatory cytokines (TNF-α), endoplasmic reticulum (ER) stress signalling proteins (GRP78, caspase-12 and GADD153) and mitogen-activated protein kinase (MAPK) family proteins (phospho-p38 MAPK and phospho-JNK) were measured by western blotting. Edaravone improved LV function in a dose-dependent manner. Central venous pressure was significantly low and LV ejection fraction and fractional shortening was significantly high in edaravone groups compared with those in the vehicle group. In addition, edaravone treatment down-regulated LV expressions of p47^phox, TNF-α, GADD153, phospho-p38 MAPK and phospho-JNK. Furthermore, the LV expressions of p67^phox, GRP78, caspase-12 and TUNEL-positive cells of rats with EAM treated with edaravone were significantly low compared with those of the vehicle group. These findings suggest that edaravone ameliorated the progression of EAM by inhibiting oxidative and ER stress and, subsequently, cardiac apoptosis.     

Wnt/β-Catenin Signaling Regulates the Proliferation and Differentiation of Mesenchymal Progenitor Cells through the p53 Pathway

Objective

Mesenchymal progenitor cells (MPCs) are found in articular cartilage from normal controls and patients with osteoarthritis (OA). Nevertheless, the molecular mechanisms of the proliferation and differentiation of these cells remain unclear. In this study, we aimed to determine the involvement of Wnt/β-catenin signaling in regulating the proliferation and differentiation of MPCs.

Methods

MPCs were isolated from the articular cartilage of normal and OA patients. Cells were sorted by immunomagnetic cell separation. Cell proliferation capacity was evaluated using the MTT assay. Toluidine blue staining and immunostaining with anti-collagen II or anti-aggrecan antibodies were used to determine the chondrogenic differentiation capabilities of MPCs. The mRNA and protein expression of target genes were examined by quantitative real-time polymerase chain reaction and Western blotting, respectively. Knock-down of p53 expression was achieved with RNA interference.

Results

Most cells isolated from the normal and OA patients were CD105⁺ and CD166⁺ positive (Normal subjects: CD105⁺/CD166⁺, 94.6%±1.1%; OA: CD105⁺/CD166⁺, 93.5%±1.1%). MPCs derived from OA subjects exhibited decreased differentiation capabilities and enhanced Wnt/β-catenin activity. Inhibition of Wnt/β-catenin signaling promoted proliferation and differentiation, whereas activation of this pathway by treatment with rWnt3a protein decreased the proliferation and differentiation of normal MPCs. Additionally, Wnt/β-catenin signaling positively regulated p53 expression, and silencing of p53 increased proliferation and differentiation of MPCs.

Conclusions

Wnt/β-catenin regulated the proliferation and differentiation of MPCs through the p53 pathway.     

Mechanical stress protects against osteoarthritis via regulation of the AMPK/NF-κB signaling pathway

Mechanical stress plays a key role in regulating cartilage degradation in osteoarthritis (OA). The aim of this study was to evaluate the effects and mechanisms of mechanical stress on articular cartilage. A total of 80 male Sprague-Dawley rats were randomly divided into eight groups (n = 10 for each group): control group (CG), OA group (OAG), and CG or OAG subjected to low-, moderate-, or high-intensity treadmill exercise (CL, CM, CH, OAL, OAM, and OAH, respectively). Chondrocytes were obtained from the knee joints of rats; they were cultured on Bioflex 6-well culture plates and subjected to different durations of cyclic tensile strain (CTS) with or without exposure to interleukin-1β (IL-1β). The results of the histological score, immunohistochemistry, enzyme-linked immunosorbent assay, and western-blot analyses indicated that there were no differences between CM and CG, but OAM showed therapeutic effects compared with OAG. However, CH and OAH experienced more cartilage damage than CG and OAG, respectively. CTS had no therapeutic effects on collagen II of normal chondrocytes, which is consistent with findings after treadmill exercise. However, CTS for 4 hr could alleviate the chondrocyte damage induced by IL-1β by activating AMP-activated protein kinase (AMPK) phosphorylation and suppressing nuclear translocation of nuclear factor (NF)-κB p65. Our findings indicate that mechanical stress had no therapeutic effects on normal articular cartilage and chondrocytes; mechanical stress only caused damage with excessive stimulation. Still, moderate biomechanical stress could reduce sensitization to the inflammatory response of articular cartilage and chondrocytes through the AMPK/NF-κB signaling pathway.     

Stimulation of α7-nAChRs coordinates autophagy and apoptosis signaling in experimental knee osteoarthritis

Osteoarthritis (OA) is the most common chronic joint disease in the elderly population. Growing evidence indicates that a balance between autophagy and apoptosis in chondrocytes plays a key role in OA’s cartilage degradation. Thus, drugs targeting the balance between apoptosis and autophagy are potential therapeutic approaches for OA treatment. In previous studies, we found that the activation of α7 nicotinic acetylcholine receptors (α7-nAChRs) alleviated monosodium iodoacetate (MIA)-induced joint degradation and osteoarthritis pain. To explore the potential functions of α7-nAChRs in autophagy and apoptosis signaling in knee OA, we compared the expression of α7-nAChRs in human knee articular cartilage tissues from normal humans and OA patients. We found that knee joint cartilage tissues of OA patients showed decreased α7-nAChRs and an imbalance between autophagy and apoptosis. Next, we observed that α7-nAChRs deficiency did not affect cartilage degradation in OA development but reversed the beneficial effects of nicotine on mechanical allodynia, cartilage degradation, and an MIA-induced switch from autophagy to apoptosis. Unlike in vivo studies, we found that primary chondrocytes from α7-nAChRs knockout (KO) mice showed decreased LC3 levels under normal conditions and were more sensitive toward MIA-induced apoptosis. Finally, we found that α7-nAChRs deficiency increased the phosphorylation of mTOR after MIA treatment, which can also be observed in OA patients’ tissues. Thus, our findings not only confirmed that nicotine alleviated MIA-induced pain behavior and cartilage degradation via stimulating the α7-nAChRs/mTOR signal pathway but found the potential role of α7-nAChRs in mediating the balance between apoptosis and autophagy.Subject terms: Autophagy, Apoptosis     

Alpha-lipoic acid attenuates methionine choline deficient diet-induced steatohepatitis in C57BL/6 mice

AimsNon-alcoholic steatohepatitis (NASH) is a liver disease that causes fat accumulation, inflammation and fibrosis. Increased oxidative stress contributes to hepatic inflammation and fibrosis by upregulation of Cytochrome P450 2E1 (CYP2E1), endoplasmic reticulum (ER) stress and mitogen-activated protein kinase (MAPK) activity. This study examined whether alpha-lipoic acid (ALA), a naturally occurring thiol antioxidant, prevents steatohepatitis through the inhibition of several pathways involved in hepatic inflammation and fibrosis.Main MethodsC57BL/6 mice were fed an MCD diet with or without ALA for 4 weeks. Liver sections from mice on control or MCD diets with or without ALA were stained with hematoxylin-eosin, oil red O, and anti-4-HNE antibody. The effects of ALA on methionine-choline deficient MCD-diet induced plasma AST and ALT as well as tissue TBARS were measured. The effects of ALA on CYP2E1 expression, ER stress, MAPK levels, and NF-κB activity in MCD diet-fed mice liver were measured by northern and western blot analysis.Key findingsDietary supplementation with ALA reduced MCD diet-induced hepatic lipid accumulation, hepatic inflammation, TBARS, 4-HNE, and plasma ALT and AST levels. These effects were associated with a reduced expression of CYP2E1 and reduced ER stress and MAPK and NF-κB activity.SignificanceTaken together, the results of the present study indicate that ALA attenuates steatohepatitis through inhibition of several pathways, and provide the possibility that ALA can be used to prevent the development and progression of non-alcoholic fatty liver disease in patients who have strong risk factors for NASH.     

Endoplasmic reticulum stress plays a role in the advanced glycation end product-induced inflammatory response in endothelial cells

Aims

Both advanced glycation end products (AGEs) and endoplasmic reticulum (ER) stress play important roles in the development of various diseases. This study aimed to clarify the consequence of AGE-induced ER stress and its underlying mechanisms in human umbilical venous endothelial cells (HUVECs).

Main methods

AGE-induced ER stress was assessed by the increased expression and activation of the ER stress marker proteins GRP78, IRE1α and JNK, which were detected using Western blot. NF-κB translocation was revealed using Western blot and immunofluorescent staining in IRE1α-knockdown HUVECs. The mechanism of AGE-induced ER stress was also explored by inhibiting the effect of reactive oxygen species (ROS) using NADPH oxidase 4 (Nox4) siRNA and the antioxidant reduced glutathione (GSH). The cellular ROS level was measured using flow cytometry.

Key findings

AGEs time- and dose-dependently enhanced the expression of GRP78 and increased the phosphorylation of IRE1α and its downstream signal JNK in HUVECs. siRNA-induced IRE1α down-regulation suppressed AGE-induced NF-κB p65 nuclear translocation. Inhibiting the ROS production using Nox4 siRNA or antagonizing ROS using GSH reduced cellular ROS level and attenuated AGE-induced GRP78 expression and IRE1α and JNK activation.

Significance

This study confirms that AGE-induced ER stress in HUVECs focuses on the ER stress-enhanced inflammatory response through JNK and NF-κB activation. It further reveals the involvement of ROS in the AGE-induced ER stress mechanism.     

Safflower yellow alleviates osteoarthritis and prevents inflammation by inhibiting PGE2 release and regulating NF-κB/SIRT1/AMPK signaling pathways

BackgroundSafflower yellow (SY) is the main active ingredient of safflower, with various pharmacological effects such as anticoagulating, antioxidant, and anti-arthritis effects.PurposeTo investigate the anti-inflammatory and chondrocyte protecting role of SY, which subsequently leads to the inhibition of cartilage degradation.MethodsRat chondrocytes were stimulated with tumor necrosis factor α (TNF-α) with or without SY treatment. Following this, CCK-8 assay was performed to detect cytotoxicity. RT-qPCR, Western blotting, and immunofluorescence staining were used to detect the gene/protein expression of typical cartilage matrix genes and related inflammatory markers. Subsequently, EdU assay was used to evaluate cell proliferation. RNA sequencing, online target prediction, and molecular docking were performed to determine the possible molecular targets and pathways.ResultsThe results showed that SY restored the TNF-α-induced up-regulation of IL-1β, PTGS2, and MMP-13 and down-regulation of COL2A1 and ACAN. Furthermore, it recovered cell proliferation by suppressing TNF-α. Gene expression profiles identified 717 differentially expressed genes (DEGs) in the cells cultured with or without SY under TNF-α stimulation. After pathway enrichment, PI3K-Akt, TNF, Cytokine-cytokine receptor interaction, NF-κB, NOD-like receptor, and Chemokine signaling pathways were notably selected to highlight NFKBIA, CCL5, CCL2, IL6, and TNF as potential targets in osteoarthritis (OA). SY inhibited TNF-α-induced activation of NF-κB and endoplasmic reticulum (ER) stress by promoting AMPK phosphorylation along with SIRT1 expression. Further, SY reduced MMP-13 expression and targeted COX-2 for decreasing PGE2 release. In addition, anterior cruciate ligament transection-induced OA was ameliorated by local administration of SY.ConclusionThese results demonstrate that SY protects chondrocytes and inhibits inflammation by regulating the NF-κB/SIRT1/AMPK pathways and ER stress, thus preventing cartilage degeneration in OA.     

Curcumin ameliorates IL-1β-induced apoptosis by activating autophagy and inhibiting the NF-κB signaling pathway in rat primary articular chondrocytes

Articular cartilage damage and chondrocyte apoptosis are common features of rheumatoid arthritis and osteoarthritis. Recently, curcumin has been reported to exhibit protective effects on degeneration in articular cartilage diseases. However, the effects and mechanisms of curcumin on articular chondrocyte injury remain to be elucidated. The aim of the present study is to investigate the chondroprotective mechanisms of curcumin on interleukin-1β (IL-1β)-induced chondrocyte apoptosis in vitro. The results revealed that IL-1β decreased cell viability and induced apoptosis in primary articular chondrocytes. Curcumin pretreatment reduced IL-1β-induced articular chondrocyte apoptosis. In addition, treatment with curcumin increased autophagy in articular chondrocytes and protected against IL-1β-induced apoptosis. The curcumin-mediated protection against IL-1β induced apoptosis was abolished when cells were treated with the autophagy inhibitor 3-methyladenine or transfected with Beclin-1 small interfering RNA. Furthermore, IL-1β stimulation significantly increased the phosphorylation levels of nuclear factor (NF)-κB p65 and glycogen synthase kinase-3β, and decreased the phosphorylation levels of β-catenin in articular chondrocytes, and these alterations to the phosphorylation levels were partly reversed by treatment with curcumin. Dual-luciferase and electrophoretic mobility shift assays demonstrated that IL-1β increased NF-κB p65 promoter activity in chondrocytes, and this was also reversed by curcumin. Pretreatment with the NF-κB inhibitor pyrrolidine dithiocarbamate enhanced the protective effects of curcumin on chondrocyte apoptosis, but Wnt/β-catenin inhibitor, XAV-939, did not exhibit this effect. Molecular docking and dynamic simulation studies results showed that curcumin could bound to RelA (p65) protein. These results indicate that curcumin may suppress IL-1β-induced chondrocyte apoptosis through activating autophagy and restraining NF-κB signaling pathway.     

Beneficial effects of angiotensin II receptor blocker,olmesartan, in limiting the cardiotoxic effect of daunorubicin in rats

Abstract

The aim was to evaluate the role of the combination of olmesartan, an angiotensin II (Ang II) receptor blocker (ARB), with daunorubicin (DNR) in reducing cardiac toxicity in rats. DNR was administered at a dose of 3 mg/kg/day every other day for 12 days. Olmesartan was administered orally every day for 12 days. Rats treated with DNR alone showed cardiac toxicity as evidenced by worsening cardiac function, elevation of malondialdehyde level in heart tissue and decreased in the level of total glutathione peroxidase activity; treatment with ARB reversed these changes. Furthermore, ARB treatment down-regulated matrix metalloproteinase-2 expression, myocardial expression of Ang II, attenuated the increased protein expressions of p67^phox and Nox4 and reduced oxidative stress-induced DNA damage evaluated by expression of 8-hydroxydeoxyguanosine. In conclusion, the result demonstrated that Ang II and oxidative stress play a key role in anthracycline-induced cardiotoxicity and that treatment with ARB will be beneficial against DNR-induced cardiotoxicity.     

Inflammation and apoptosis in aortic tissues of aged type II diabetes: Amelioration with α-lipoic acid through phosphatidylinositol 3-kinase/Akt- dependent mechanism

AimsEndothelial dysfunction is a key triggering event in the development of cardiovascular diseases and the current study explored this phenomenon in the context of inflammation, apoptosis, reactive oxygen species (ROS) and the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway during chronic diabetes.Main methodsα-Lipoic acid (ALA) and wortmannin (WM) were chronically administered to aged Goto Kakizaki (GK) rats, a genetic model of non-obese type II diabetes. Key indices of inflammation, apoptosis and oxidative stress were assessed using western blotting, real-time PCR and immunofluoresence-based techniques.Key findingsA chronic inflammation (e.g., increased mRNA/protein levels of TNF-α, ICAM, fractalkine, CD-68, myeloperoxidase) in connection with increased caspase-based apoptotic cell death and heightened state of oxidative stress (HSOS)– appear to exist in diabetic cardiovascular tissues. An assessment of NF-κB dynamics in aged diabetic vessels revealed not only a marked increase in cytosolic phosphorylated levels of IκB-α, NIK, IKK but also an enhancement in nuclear localization of p65 concomitantly with augmented NF-κB-DNA binding activity. Most of the aforementioned cardiovascular-based diabetic abnormalities including reduced activities of PI3K and Akt kinase were ameliorated following chronic ALA therapy. WM, given to GK rats negated the anti-inflammatory and anti-apoptotic actions of ALA.SignificanceOur data highlight a unifying mechanism whereby HSOS through an induction of NF-κB activity together with an impairment in PI3K/Akt pathway favors pro-inflammatory/pro-apoptotic diabetic vascular milieu that culminate in the onset of endothelial dysfunction, a phenomenon which appears to be amenable to treatment with antioxidants and/or PI3/Akt mimetics (e.g., ALA).     

Mechanism of endoplasmic reticulum stress-induced vascular endothelial dysfunction

Background

We recently reported that ER stress plays a key role in vascular endothelial dysfunction during hypertension. In this study we aimed to elucidate the mechanisms by which ER stress induction and oxidative stress impair vascular endothelial function.

Methodology/principal findings

We conducted in vitro studies with primary endothelial cells from coronary arteries stimulated with tunicamycin, 1 μg/mL, in the presence or absence of two ER stress inhibitors: tauroursodeoxycholic acid (Tudca), 500 μg/mL, and 4-phenylbutyric acid (PBA), 5 mM. ER stress induction was assessed by enhanced phosphorylation of PERK and eIF2α, and increased expression of CHOP, ATF6 and Grp78/Bip. The ER stress induction increased p38 MAPK phosphorylation, Nox2/4 mRNA levels and NADPH oxidase activity, and decreased eNOS promoter activity, eNOS expression and phosphorylation, and nitrite levels. Interestingly, the inhibition of p38 MAPK pathway reduced CHOP and Bip expressions enhanced by tunicamycin and restored eNOS promoter activation as well as phosphorylation. To study the effects of ER stress induction in vivo, we used C57BL/6J mice and p47phox^−/− mice injected with tunicamycin or saline. The ER stress induction in mice significantly impaired vascular endothelium-dependent and independent relaxation in C57BL/6J mice compared with p47phox^−/− mice indicating NADPH oxidase activity as an intermediate for ER stress in vascular endothelial dysfunction.

Conclusion/significance

We conclude that chemically induced ER stress leads to a downstream enhancement of p38 MAPK and oxidative stress causing vascular endothelial dysfunction. Our results indicate that inhibition of ER stress could be a novel therapeutic strategy to attenuate vascular dysfunction during cardiovascular diseases.     

Zinc protects chondrocytes from monosodium iodoacetate-induced damage by enhancing ATP and mitophagy

Osteoarthritis (OA) is characterized with articular cartilage degradation, and monosodium iodoacetate (MIA)-treated chondrocyte is the most commonly used model for mimicking OA progression. Zinc protects chondrocytes from MIA-induced damage. Here, we explored the protective effects of 25 μM zinc on 5 μM MIA-treated SW1353 cells (human chondrosarcoma cell line) through the analysis of energy metabolism- and autophagy-related parameters. We found that the exposure of SW1353 cells to MIA decreased ATP levels, expression of glycolysis-related proteins, including glucose transporter 1, hexokinase 2, and pyruvate dehydrogenase E1 component subunit alpha, and the levels of mitochondrial complex I, II, IV, and V subunits of the oxidative phosphorylation pathway. MIA treatment also decreased the expression of autophagy-related proteins, including autophagic elongation protein 5 (ATG5), ATG7, and microtubule-associated protein 1A/1B light chain 3B (LC3-II) and mitophagy-related proteins, including phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1), ubiquitin, and p62. These results indicate that MIA interferes with energy metabolism and the autophagic clearance of dysfunctional mitochondria (so called mitophagy). Interestingly, zinc exposure could almost completely reverse the effects of MIA, suggesting its potential protective role against OA progression.     

Multifactor dimensionality reduction reveals a strong gene–gene interaction between STC1 and COL11A1 genes as a possible risk factor of knee osteoarthritis

Articular cartilage is an avascular tissue with a structure that allows it to support and cushion the overload of the surfaces in contact. It maintains its metabolic functions due to the contribution of different signaling pathways. However, several factors play a role in its deterioration, allowing to the development of osteoarthritis (OA), and one of the major factors is genetic. Our goal was to identify gene–gene interactions (epistasis) between five signaling pathways involved in the articular cartilage metabolism as possible indicators of OA risk. We applied the Multifactor-Dimensionality Reduction (MDR) method to identify and characterize the epistasis between 115 SNPs located in 73 genes related to HIF-1α, Wnt/β-catenin, cartilage extracellular matrix metabolism, oxidative stress, and uric acid transporters. Ninety three patients diagnosed with primary knee OA and 150 healthy controls were included in the study. Genotyping was performed with the OpenArray system, the statistical analysis was carried out with the STATA software v14, and epistasis was analyzed with the MDR software v3.0.2. The MDR analysis revealed that the best interaction model was between polymorphisms rs17786744 of the STC1 gene and rs2615977 of the COL11A1 gene, with an entropy value of 4.44%, CVC 8/10, OR 5.60, 95% CI 3.27–9.59, p?<?0.0001. Under this interaction model, we identified high and low risk genotypes involved in OA development. Our results suggest complex interactions between STC1 and COL11A1 genes that might have an impact on genetic susceptibility to develop OA. Further studies are required to confirm it.

     

Role for the first SH3 domain of p67 in activation of superoxide-producing NADPH oxidases

The membrane-bound NADPH oxidase in phagocytes, gp91^phox (a.k.a. Nox2), produces superoxide, a precursor of microbicidal oxidants, thereby playing a crucial role in host defense. Activation of gp91^phox/Nox2 requires assembly with the cytosolic proteins p67^phox and p47^phox, each containing two SH3 domains. Although the C-terminal SH3 domain of p67^phox is responsible for binding to p47^phox, little is known about the role for the first (N-terminal) SH3 domain [SH3(N)]. Here we show that truncation of p67^phox-SH3(N), but not substitution of arginine for the invariant residue Trp-277 in SH3(N), results in an impaired activation of gp91^phox/Nox2. The impairment is overcome by higher expression of an SH3(N)-defective p67^phox in cells, suggesting that SH3(N) primarily increases the affinity of p67^phox for the oxidase complex. On the other hand, p67^phox-SH3(N) is not involved in activation of Nox1 and Nox3, closely-related homologues of gp91^phox/Nox2. Thus p67^phox-SH3(N) specifically functions in gp91^phox/Nox2 activation probably via facilitating oxidase assembly.