Lipid metabolism and osteoarthritis: lessons from atherosclerosis

Osteoarthritis (OA) is an age-related degenerative disease comprising the main reason of handicap in the Western world. Interestingly, to date, there are neither available biomarkers for early diagnosis of the disease nor any effective therapy other than symptomatic treatment and joint replacement surgery. OA has long been associated with obesity, mainly due to mechanical overload exerted on the joints. Recent studies however, point to the direction that OA is a metabolic disease, as it also involves non-weight bearing joints. In fact, altered lipid metabolism may be the underlying cause. First, adipokines have been shown to be key regulators of OA pathogenesis. Second, epidemiological studies have shown serum cholesterol to be a risk factor for OA development. Third, lipid deposition in the joint is observed at the early stages of OA before the occurrence of histological changes. Fourth, proteomic analyses have shown an important connection between OA and lipid metabolism. Finally, recent gene expression studies reveal a deregulation of cholesterol influx and efflux and in the expression of lipid metabolism-related genes. Interestingly, lipids and lipid metabolism are known to be implicated in the development and progression of another age-related degenerative disease, atherosclerosis (ATH). Thus, although it is tempting to speculate that the osteoarthritic chondrocyte has been transformed to foam cell, it has not been proven yet. However, this may be an intriguing theory linking ATH and OA, which may open new avenues to novel therapeutic interventions for OA taking advantage of previous knowledge from ATH.     

Detection and characterization of interleukin-6 gene variants in Canis familiaris: association studies with periodontal disease

Periodontal disease (PD) is the most common inflammatory disease of the oral cavity of domestic carnivores. In Human Medicine molecular genetics research showed that several genes play a role in the predisposition and progression of this complex disease, primarily through the regulation of inflammatory mediators, but the exactly mechanisms are poorly understood. This study aims to contribute to the characterization of the genetic basis of PD in the dog, a classically accepted model in Periodontology. We searched for genetic variations in the interleukin-6 (IL6) gene, in order to verify its association with PD in a case-control study including 25 dogs in the PD case group and 45 dogs in the control group. We indentified and characterized three new genetic variations in IL6 gene. No statistically significant differences were detected between the control and PD cases groups. Our results do not support an evidence for a major role contribution of these variants in the susceptibility to PD in the analyzed population. Nevertheless, the sequence variant I/5_g.105 G > A leads to an amino acid change (arginine to glutamine) and was predicted to be possibly damaging to the IL6 protein. A larger cohort and functional studies would be of extreme importance in a near future to understand the possible role of IL6 variants in this disease.     

Safety issues of Lactobacillus bulgaricus with respect to human gelatinases in vitro

In oral medicine and dentistry probiotics have shown promising results in controlling dental diseases and yeast infections. This study was made to investigate the effect of eight strains of Lactobacillus bulgaricus and their effects on human matrix metalloproteinase-9 (MMP-9). The hypothesis was that these bacteria used in yoghurt production for centuries are not proteolytic and thus can be safely used in the development of probiotic preparations. Bacterial cell fractions and supernatant specimens were prepared and studied with gelatinase zymography and MMP-9 activation was assessed by immunoblotting. The effect of synthetic MMP inhibitors and a serine protease inhibitor (Pefabloc) on bacterial proteinases was studied with zymography. The results showed very low gelatinolytic activity. There was a slight difference between the supernatant and cell fractions so that the supernatant specimens produced weak gelatinolytic bands in zymography while hardly anything was seen in the cell fraction series. The tested synthetic MMP inhibitors and Pefabloc did not affect the proteolytic activity of the lactobacilli strains. The lactobacilli did not seem to induce the conversion of proMMP-9 to its active form. Consequently, our study hypothesis was confirmed and the studied Lactobacillus strains are not likely to degrade host tissue components.     

An in situ hybridization study of MMP-2, -9, -13, -14, TIMP-1, and -2 mRNA in fetal mouse mandibular condylar cartilage as compared with limb bud cartilage

The main purpose of this in situ hybridization study was to investigate MMPs and TIMPs mRNA expression in developing mandibular condylar cartilage and limb bud cartilage. At E14.0, MMP-2, -14, TIMP-1 and -2 mRNAs were expressed in the periosteum of mandibular bone, and in the condylar anlage. At E15.0 MMP-2, -14, TIMP-1 and -2 mRNAs were expressed in the perichondrium of newly formed condylar cartilage and the periosteum of developing bone collar, whereas, expression of MMP-14 and TIMP-1 mRNAs were restricted to the inner layer of the periosteum/perichondrium. This expression patterns continued until E18.0. Further, from E13.0 to 14.0, in the developing tibial cartilage, MMP-2, -14, and TIMP-2 mRNAs were expressed in the periosteum/perichondrium, but weak MMP-14 and no TIMP-1 mRNA expression was recognized in the perichondrium. These results confirmed that the perichondrium of condylar cartilage has characteristics of periosteum, and suggested that MMPs and/or TIMPs are more actively involved in the development of condylar (secondary) cartilage than tibial (primary) cartilage. MMP-9-positive cells were observed in the bone collar of both types of cartilage, and they were consistent with osteoclasts/chondroclasts. MMP-13 mRNA expression was restricted to the chondrocytes of the lower hypertrophic cell zone in tibial cartilage at E14.0, indicating MMP-13 can be used as a marker for lower hypertrophic cell zone. It was also expressed in chondrocytes of newly formed condylar cartilage at E15.0, and continuously expressed in the lower hypertrophic cell zone until E18.0. These results confirmed that progenitor cells of condylar cartilage are rapidly differentiated into hypertrophic chondrocytes, which is a unique structural feature of secondary cartilage different from that of primary cartilage.     

Dynamic interplay between breast cancer cells and normal endothelium mediates the expression of matrix macromolecules,proteasome activity and functional properties of endothelial cells

Background

Breast cancer–endothelium interactions provide regulatory signals facilitating tumor progression. The endothelial cells have so far been mainly viewed in the context of tumor perfusion and relatively little is known regarding the effects of such paracrine interactions on the expression of extracellular matrix (ECM), proteasome activity and properties of endothelial cells.

Methods

To address the effects of breast cancer cell (BCC) lines MDA-MB-231 and MCF-7 on the endothelial cells, two cell culture models were utilized; one involves endothelial cell culture in the presence of BCCs-derived conditioned media (CM) and the other co-culture of both cell populations in a Transwell system. Real-time PCR was utilized to evaluate gene expression, an immunofluorescence assay for proteasome activity, and functional assays (migration, adhesion and invasion) and immunofluorescence microscopy for cell integrity and properties.

Results

BCC-CM decreases the cell migration of HUVEC. Adhesion and invasion of BCCs are favored by HUVEC and HUVEC-CM. HA levels and the expression of CD44 and HA synthase-2 by HUVEC are substantially upregulated in both cell culture approaches. Adhesion molecules, ICAM-1 and VCAM-1, are also highly upregulated, whereas MT1-MMP and MMP-2 expressions are significantly downregulated in both culture systems. Notably, the expression and activity of the proteasome β5 subunit are increased, especially by the action of MDA-MB-231-CM on HUVEC.

Conclusions and general significance

BCCs significantly alter the expression of matrix macromolecules, proteasome activity and functional properties of endothelial cells. Deep understanding of such paracrine interactions will help to design novel drugs targeting breast cancer at the ECM level. This article is part of a Special Issue entitled Matrix-mediated cell behaviour and properties.     

EMMPRIN/CD147-encriched membrane vesicles released from malignant human testicular germ cells increase MMP production through tumor–stroma interaction

Background

Elevated levels of EMMPRIN/CD147 in cancer tissues have been correlated with tumor progression but the regulation of its expression is not yet understood. Here, the regulation of EMMPRIN expression was investigated in testicular germ cell tumor (TGCTs) cell lines.

Methods

EMMPRIN expression in seminoma JKT-1 and embryonal carcinoma NT2/D1 cell lines was determined by Western blot, immunofluorescence and qRT-PCR. Membrane vesicles (MVs) secreted from these cells, treated or not with EMMPRIN siRNA, were isolated by differential centrifugations of their conditioned medium. MMP-2 was analyzed by zymography and qRT-PCR.

Results

The more aggressive embryonic carcinoma NT2/D1 cells expressed more EMMPRIN mRNA than the seminoma JKT-1 cells, but surprisingly contained less EMMPRIN protein, as determined by immunoblotting and immunostaining. The protein/mRNA discrepancy was not due to accelerated protein degradation in NT2/D1 cells, but by the secretion of EMMPRIN within MVs, as the vesicles released from NT2/D1 contained considerably more EMMPRIN than those released from JKT-1. EMMPRIN-containing MVs obtained from NT2/D1, but not from EMMPRIN-siRNA treated NT2/D1, increased MMP-2 production in fibroblasts to a greater extent than those from JKT-1 cells.

Conclusion and general significance

The data presented show that the more aggressive embryonic carcinoma cells synthesize more EMMPRIN than seminoma cells, but which they preferentially target to secreted MVs, unlike seminoma cells which retain EMMPRIN within the cell membrane. This cellular event points to a mechanism by which EMMPRIN expressed by malignant testicular cells can exert its MMP inducing effect on distant cells within the tumor microenvironment to promote tumor invasion. This article is part of a Special Issue entitled Matrix-mediated cell behaviour and properties.     

Genetic predisposition to left ventricular dysfunction: a multigenic and multi-analytical approach

Background

Left ventricular dysfunction (LVD) is a complex, multifactorial condition, caused by mechanical, neurohormonal, and genetic factors. We have previously observed association of renin–angiotensin–aldosterone system (RAAS), matrix metalloproteinases (MMPs) and inflammatory pathway genes with LVD. Therefore the present study was undertaken to identify the combination of genetic variants and their possible interactions contributing towards genetic susceptibility to LVD in the background of coronary artery disease (CAD).

Methods and results

The study included 230 healthy controls and 510 consecutive patients with angiographically confirmed CAD. Among them, 162 with reduced left ventricle ejection fraction (LVEF ≤ 45%) were categorized as having LVD. We analyzed 11 polymorphisms of RAAS, MMPs and inflammatory pathways. Single locus analysis showed that AT1 A1166C (p value < 0.001; OR = 3.67), MMP9 R668Q (p value = 0.007; OR = 3.48) and NFKB1-94 ATTG ins/del (p value = 0.013; OR = 2.01) polymorphisms were independently associated with LVD when compared with both non-LVD patients and healthy controls. High-order gene–gene interaction analysis, using classification and regression tree (CART) and multifactor dimensionality reduction (MDR) revealed that AT1 A1166C and NFKB1-94 ATTG ins/del polymorphisms jointly increased the risk of LVD to great extent (p-value = 0.001; OR = 8.55) and best four-factor interaction model consisted of AT1 A1166C, MMP7 A-181G, MMP9 R668Q and NFKB1-94 ATTG ins/del polymorphisms with testing accuracy of 0.566 and cross validation consistency (CVC) = 9/10 (permutation p < 0.001) showed increased risk for LVD respectively.

Conclusion

AT1 A1166C independently and in combination with MMP9 R668Q and NFKB1-94 ATTG ins/del polymorphisms plays important role in conferring genetic susceptibility to LVD in CAD patients.     

Curcumin inhibits the invasion of thyroid cancer cells via down-regulation of PI3K/Akt signaling pathway

The objectives of this study were to evaluate the in vitro anti-tumor (human thyroid cancer cell lines) potential of curcumin and to elucidate its molecular mechanisms. Here, we investigated the effects of curcumin on the cell viability, apoptosis, migration and invasion of human thyroid cancer cell lines FTC133. We also investigated the effects of curcumin on PI3K, p-Akt, MMP1/7, and COX-2 protein expressions using Western blot. Results showed that curcumin inhibited growth, cell migration and invasion in FTC133, and promoted its apoptosis. Western blot assay data demonstrated that curcumin inhibited phosphorylation of PI3K and Akt signaling pathways and subsequently attenuated MMP1/7 and COX-2 protein expressions in FTC133. In conclusion, curcumin suppresses FTC133 cell invasion and migration by inhibiting PI3K and Akt signaling pathways. Therefore, curcumin produces anti-metastatic activity in FTC133 cells.     

Endometrial stromal cells and decidualized stromal cells: Origins,transformation and functions

Decidualization of endometrium, which is characterized by endometrial stromal cell (ESC) decidualization, vascular reconstruction, immune cell recruitment, and plentiful molecule production, is a crucial step for uterus to become receptive for embryo. When implantation takes place, ESCs surround and directly interact with embryo. Decidualized stromal cells (DSCs) are of great importance in endometrial decidualization, having a broad function in regulating immune activity and vascular remodeling of uterus. DSCs are shown to have a higher metabolic level and looser cytoskeleton than ESCs. What's the origin of ESCs and how ESCs successfully transform into DSCs had puzzled scientists in the last decades. Breakthrough had been achieved recently, and many studies had elucidated some of the characters and functions of DSCs. However, several questions still remain unclear. This paper reviews current understanding of where ESCs come from and how ESCs differentiate into DSCs, summarizes some characters and functions of DSCs, analyzes current studies and their limitations and points out research areas that need further investigation.     

Candesartan antagonizes pressure overload-evoked cardiac remodeling through Smad7 gene-dependent MMP-9 suppression

The present study was designed to investigate the underlying molecular mechanism for Angiotensin II type 1 receptor blockers (ARBs) mediated cardio-protection against pressure overload-induced cardiac remodeling with a focus on Smad7. ROCK-1, Smad3 and fibronectin expressions were increased in male C57BL/6 mice underwent transverse aortic constriction (TAC) for 2weeks. Treatment with Candesartan (2mg/kg per day) could effectively downregulate Smad3 and fibronectin accompanied by upregulating of Smad7. Further data showed that Candesartan inhibited TGF-β1 signal-induced epithelial-to-mesenchymal transition (EMT) through attenuating matrix metalloproteinases (MMP-9), such effect was abolished by knocking-down Smad7. Moreover, TAC for 2weeks caused increased collagen deposition, thickness of left ventricular anterior and posterior wall at end-diastole (LVAWD and LVPWD) and LVEF% reduction, which were reversed by treatment with Candesartan, but failed after knocking-down Smad7. In addition, LV dP/dt(max) and dP/dt(min) were increased by TAC for 2weeks, and treatment with Candesartan or Nifedipine effectively depressed the high levels of dP/dt(min) induced by TAC. However, only Candesartan-mediated protective role in improving cardiac function was suppressed by tail-vein injection of Smad7 siRNA. This study uncovered a novel role for ARBs in preventing pressure overload-induced cardiac remodeling via Smad7 upregulation, which suppressed MMP-9 expression and TGF-β1 signal-mediated EMT progress.