Assunta Pozzuoli

High-density lipoproteins downregulate CCL2 production in human fibroblast-like synoviocytes stimulated by urate crystals

IntroductionTo investigate whether monosodium urate (MSU) crystals induce the production of CCL2 (monocyte chemoattractant protein-1; MCP-1) in human fibroblast-like synoviocytes (FLS) and whether this mechanism would be affected by high-density lipoproteins (HDL).MethodsHuman FLS isolated from synovial tissue explants were stimulated with MSU crystals (0.01 to 0.5 mg/ml) or interleukin (IL)-1β (10 pg/ml) in the presence or absence of HDL (50 and 100 μg/ml). The production and expression of CCL2 was evaluated with ELISA, confocal microscopy, immunofluorescence microscopy, chemotaxis assay, and real-time quantitative PCR.ResultsExposure of FLS to MSU crystals induced CCL2 accumulation in culture medium in a dose- and time-dependent manner, reaching a plateau at 50 to 75 μg/ml MSU crystals and 20 to 24 hours. Although low, the induced CCL2 levels were sufficient to trigger mononuclear cell migration. In resting FLS, CCL2 was localized in small cytoplasmic vesicles whose number diminished with MSU crystal stimulation. Concomitantly, MSU crystals triggered the induction of CCL2 mRNA expression. All these processes were inhibited by HDL, which cause a 50% decrease in CCL2 mRNA levels and a dose-dependent inhibition of the release of CCL2. Similar results were obtained when FLS were pretreated with HDL and washed before activation by MSU crystals or IL-1β, suggesting a direct effect of HDL on the FLS activation state.ConclusionsThe present results demonstrate that MSU crystals induce FLS to release CCL2 that is stored in vesicles in resting conditions. This mechanism is inhibited by HDL, which may limit the inflammatory process by diminishing CCL2 production and, in turn, monocytes/macrophages recruitment in joints. This study confirms the antiinflammatory functions of HDL, which might play a part in the limitation of acute gout attack.

Tryptophan Catabolism in Synovial Fluid of Various Arthropathies and its Relationship with Inflammatory Cytokines

Synovial fluids (SF) from patients with rheumatoid arthritis (RA), psoriatic arthritis (PsA), gout, and osteoarthritis (OA) were investigated for the levels of interleukin (IL)-1 beta, IL-6 and IL-8, tryptophan (Trp) and indoleamine 2,3-dioxygenase (IDO) activity. Significant differences exist in the levels of IL-1 beta between inflammatory arthritides RA, PsA and gout and non inflammatory arthritis, such as OA. The highest concentration of IL-1 beta was found in RA, that showed high levels also of IL-6 and IL-8. In the same disease we also found the highest IDO activity and the lowest Trp concentration. In addition, IDO activity seems to be related with the decrease in Trp, as demonstrated by the inverse correlation found between these two substances in the SF of all patients.

Activated T cells sustain myeloid-derived suppressor cell-mediated immune suppression

The expansion of myeloid derived suppressor cells (MDSCs), a suppressive population able to hamper the immune response against cancer, correlates with tumor progression and overall survival in several cancer types. We have previously shown that MDSCs can be induced in vitro from precursors present in the bone marrow and observed that these cells are able to actively proliferate in the presence of activated T cells, whose activation level is critical to drive the suppressive activity of MDSCs. Here we investigated at molecular level the mechanisms involved in the interplay between MDSCs and activated T cells. We found that activated T cells secrete IL-10 following interaction with MDSCs which, in turn, activates STAT3 phosphorylation on MDSCs then leading to B7-H1 expression. We also demonstrated that B7-H1+ MDSCs are responsible for immune suppression through a mechanism involving ARG-1 and IDO expression. Finally, we show that the expression of ligands B7-H1 and MHC class II both on in vitro-induced MDSCs and on MDSCs in the tumor microenvironment of cancer patients is paralleled by an increased expression of their respective receptors PD-1 and LAG-3 on T cells, two inhibitory molecules associated with T cell dysfunction. These findings highlight key molecules and interactions responsible for the extensive cross-talk between MDSCs and activated T cells that are at the basis of immune suppression.

The Lumbar Interspinous Ligaments in Humans: Anatomical Study and Review of the Literature

In textbooks of human anatomy and atlases of spinal surgery, the lumbar interspinous ligaments are described and illustrated in various and often conflicting ways. Thirty-four lumbar vertebral columns (19 males, 15 females, range 17–92 years old), sampled at autopsy, were studied by dissection and macroscopic analysis. The orientation of the fibre bundles was evaluated in the sagittal plane by tracing a reference line parallel to the cranial border of the spinous process, and the degree of bundle tilting with reference to this line was measured in the ventral, middle and dorsal parts of the ligament. Histological and immunohistochemical (anti-S100) studies of the ligaments, on sagittal, frontal, and transversal planes, were also carried out. In vivo radiological validation was performed in 25 patients by MR and CT imaging. Macroscopically, the interspinous ligaments are consistently composed of bundles of fibres arranged in a characteristic pattern. Together, they present an oblique orientation from anterior to posterior in a caudal-cranial direction. In their ventral part the bundles form a slight curve with a posterior-inferior concavity. In the middle part, the bundles are thicker and arranged in an italic S shape with a mean degree of tilting of 52 ± 14, 21 ± 12 and 50 ± 17 in the anterior, intermediate and posterior subzones of the L2–L3 ligaments and of 32 ± 14, 11 ± 9 and 18 ± 11 in the anterior, intermediate and posterior subzones of the L4–L5 level. The dorsal part consists of obliquely ascending bundles of fibres mostly converging in the supraspinous ligament. Histologically, these ligaments are mainly composed of collagen fibres, whereas the elastic fibres are ubiquitous, although mostly concentrated in the ventral part, which is closely linked to the yellow ligament. The interspinous ligament is well supplied by small blood vessels and sensory nerves, the latter particularly in its dorsal part and on its lateral surfaces. According to collected data, the structure of the interspinous ligaments reflects their function, i.e., the italic S-shaped course of the fibrous bundles represents an available reserve of length in a poorly elastic ligament.

In vitro response of osteoarthritic chondrocytes and fibroblast-like synoviocytes to a 500–730 kDa hyaluronan amide derivative†

The aim of this study was to compare the effects of native hyaluronan (HA) with that of its hexadecylamide derivative (HYADD) on proliferation of fibroblast-like synoviocytes (FLS) and chondrocytes. The production of inflammatory and anti-inflammatory cytokines was also analyzed in FLS cultures. The proliferation of osteoarthritis (OA) chondrocytes was enhanced when cells were treated with 0.5-1.5 mg mL(-1) of HA or HYADD®4-G. This effect was completely suppressed by the anti-CD44 antibody. At 0.5 to 1 mg mL(-1) , HA and HYADD®4-G did not influence the proliferation of normal or pathological FLS; however, at the higher concentration (1.5 mg mL(-1) ), HYADD®4-G did significantly inhibit cell proliferation. As to effects on inflammation, a significant increase in the expression of the IL-10 gene was observed when FLS were pretreated with tumor necrosis factor alpha and then cultured in the presence of 0.5 mg mL(-1) HYADD® 4-G or HA. The effects of HA derivatives on FLS proliferation and production of anti-inflammatory cytokines indicate that they may be of therapeutic benefit in OA. The longer residence time in the joint cavity, the increased viscoelasticity, and the anti-inflammatory potential of HYADD®4-G make it a better candidate than native HA for OA therapy.

Effects of three therapeutic regimens on postmenopausal bone loss in oophorectomized women

Abstract Seventy-five patients, who underwent hysterectomy and bilateral oophorectomy for nonmalignant disease, were randomly allocated into four groups and treated with three different therapeutic regimens in an open-label prospective study. Group A was given nasal spray salmon calcitonin 100 IU/d, group B was given transdermal estradiol 50 μg twice a week, group C was given calcium 500 mg/d plus 1α-hydroxyvitamin D 3 (vitamin D) 25 μg/d, and group D was the control group. All groups showed significant bone loss during the 36-month study; however, in the estrogen-treated group, bone loss was significantly lower than that observed in the other groups. Bone GLA protein increased in all groups, but the increase was less in the estrogen-treated group. Calcitonin and calcium plus vitamin D were ineffective in protecting women from postoophorectomy bone loss and from an increase of bone turnover. Estrogen replacement therapy was the most effective of the three therapeutic regimens tested in this study, even if this regimen was unable to completely stop postoophorectomy bone loss.

Management of the subscapularis contracture during shoulder arthroplasty for primary glenohumeral arthritis

AimTo evaluate the safety and effectiveness of a particular subscapularis release in shoulder arthroplasty for primary glenohumeral arthritis.Materials and methodsTwenty-eight patients (19F, 9M) underwent shoulder arthroplasty for primary glenohumeral arthritis. Preoperative average Constant Score (CS) was 31.2 points (range 14–52), active anterior elevation (AAE) 92° (30–100°) and active external rotation (AER) 11° (−40 to 20°). During arthroplasty for subscapularis contracture, patients underwent subscapularis release freeing the superior tubular tendon (STT) with a section of the coracohumeral ligament (CHL) and the superior glenohumeral ligament (SGHL) and a deep release consisting of a section of the middle glenohumeral ligament (MGHL), very close to the glenoid labrum, and the inferior glenohumeral ligament (IGHL). An anatomic study was performed on 13 cadavers, verifying the structure of subscapularis tendon and its relationship with the capsule, the surrounding ligaments and the axillary nerve. Moreover, after having placed traction sutures on the subscapularis tendon, its lengthening was measured after STT release alone and after STT and deep release. The complete absence of neurological and vascular lesions was also verified.ResultsAverage follow-up: 2.9 years. Postoperative mean CS was 70.5 (p〈0.005), with an absolute gain of 39.1. AAE increased from 92° to 142° (p=0.001) while AER increased from 8° to 48° (p=0.002). At the last follow-up, 19 patients (67.8%) were very satisfied, 5 patients (17.8%) were satisfied, 3 patients (10.7%) partially satisfied and 1 patient (3.5%) unsatisfied. In the anatomic control, the average lengthening of subscapularis tendon was 0.9 cm after STT release alone and 2.5 cm after STT and deep release. No vascular and neurological lesions were observed.ConclusionsThe subscapularis release during shoulder arthroplasty is extremely important to obtain the proper balance between anterior and posterior soft tissues and to achieve an optimal range of motion and joint stability. An adequate anatomical dissection could give good tendon mobilisation and lengthening, necessary for a good repair, and lead to a recovery of the range of motion, particularly for external rotation.

Psoriatic arthritis exacerbated by Salmonella infection.

Abstract: Psoriatic arthritis (PsA) is an inflammatory joint disease in which environmental factors, particularly trauma and infections, are thought to play an important role. The authors describe the case of a patient with a mild and long-untreated form of PsA which was severely exacerbated by Salmonella typhimurium infection. This case confirms the importance of infectious agents in the occurrence and course of PsA.

Conditioned media from human osteoarthritic synovium induces inflammation in a synoviocyte cell line

ABSTRACT Aim: Osteoarthritis (OA) is a whole joint pathology involving cartilage, synovial membrane, meniscus, subchondral bone, and infrapatellar fat pad (IFP). Synovitis has been widely documented in OA suggesting its important role in pathogenesis. The aim of this study was to investigate the role of different joint tissues in promoting synovitis. Materials and methods: Conditioned media (CM) from cartilage, synovial membrane, meniscus, and IFP were generated from tissues of five patients undergoing total knee replacement and used to stimulate a human fibroblast-like synoviocytes cell line (K4IM). Cytokines, chemokines, and metalloproteases release was analyzed in all CM by Bio-Plex Assay and sulfated glycosaminoglycan (GAG) content by dimethylmethylene blue assay. Gene expression of several markers was evaluated by real-time PCR in K4IM cells stimulated with the CM obtained from joint tissues. Results: CM from all tissues produced high levels of IL-6, IL-8, and CCL2. CCL21, MMP-3, and −13 levels were detected in all CM except IFP. MMP-10 was present only in CM of cartilage and synovial tissues. IL-1β, IL-15, TNF-α, CCL5, and CCL19 were undetectable. However, only K4IM cells stimulated by the CM from OA synovium showed an increase of IL-6, CXCL-8, CCL21, MMP10, and IL-1β expression. Conclusion: Our study showed that K4IM might be a suitable in vitro model for evaluating different cellular pathways in OA studies. Importantly, we demonstrated that in OA, all joint tissues might be involved in the progression of synovitis with a predominant role of synovial membrane itself compared to the other joint tissues.

The challenging management of a delayed union midshaft clavicle fracture complicated by an acute pseudoaneurysm of the subclavian artery in a superelderly diabetic patient

Clavicle fractures are among the most common fractures, accounting for 2.6–4% of all adult fractures and for 35–44% of those of the shoulder girdle [1]. Up to 80% occur at the midshaft. Nowadays, the majority of these fractures tend to be treated non-operatively, even when displaced, using an arm supporting sling or a “figure-of-eight” bandage, with good clinical outcomes and an acceptable rate of nonunion. On the contrary, operative treatment is commonly performed in cases of open fractures, skin tenting with the potential for progression to open fracture, “floating shoulder,” and associated acute neurovascular injuries. Despite the proximity of the clavicle to the subclavian vessels, vascular complications in closed clavicle fractures are uncommon, with an incidence of 0.4% [2]. Nevertheless, their prompt diagnosis and proper knowledge is essential because of the high morbidity and mortality rates associated. This report describes the clinical, diagnostic, and therapeutic approaches to address an acute subclavian artery pseudoaneurysm caused by a closed displaced clavicle fracture, complicated by delayed union, in a comorbid octogenarian patient. Presentation of case