A correlation between knee cartilage degradation observed by arthroscopy and synovial proteinases activities

OBJECTIVE: A novel study has been carried out to characterize the amount and activity levels of metalloproteinases (i.e., MMP-1, MMP-2, MMP-3, MMP-8, MMP-9 and MMP-13) and of their inhibitors (i.e., TIMP-1 and TIMP-2) in synovial fluid from patients (n = 56) with different degrees of either chondral lesions or knee arthritis identified and classified by arthroscopy. DESIGN AND METHODS: Zymographies, Western blotting and ELISA tests have been used to correlate the disease stage, as determined by arthroscopy, and both the amount and the activation state of different MMPs and of their inhibitors. RESULTS: Analysis of data obtained demonstrates that the degree of cartilage degradation, as seen by arthroscopy, is strictly related to the activity of some synovial MMPs, in particular MMP-2 and MMP-13 and on reduced inhibitory effect of MMP-2 by TIMP-2; in addition, a serine protease weighing about 125 kDa appears only in patients with severe cartilage degradation, i.e., with knee arthritis. CONCLUSIONS: On the whole, this is the first study in which an analysis of synovial MMPs/other proteinases activity and TIMPs has been strictly related to arthroscopy results in patients with different degrees of osteoarthritis. Results indicate that an imbalance between specific MMP activities and the amount of TIMPs and of its inhibitory efficiency is crucial for the disease evolution and it is related to the disease stage.     

Novel Synthetic,Salt-Resistant Analogs of Human Beta-Defensins 1 and 3 Endowed with Enhanced Antimicrobial Activity

Human beta-defensins (hBDs) are antimicrobial peptides of human innate immunity. The antibacterial activities of hBDs 1, 2, and 4 but not the activity of hBD3 are impaired by high salt levels. We have designed and synthesized seven novel hBD analogs, constituted by different domains of hBD1 (which is constitutively expressed in humans) and of hBD3 (which is induced by microorganisms and inflammatory factors in humans), that would maintain and potentially increase the wild-type antimicrobial activities and be salt resistant. We have compared the antibacterial, antiviral, and chemotactic activities of the analogs with those of hBD1 and hBD3. We show that the hBD1 internal region and the hBD3 C-terminal region are critical for antibacterial activity also at high salt concentrations, whereas deletion of the N-terminal region of hBD3 results in an increase in antibacterial activity. All analogs inhibited herpes simplex virus; antiviral activity was enhanced by the hBD1 internal region and the hBD3 C-terminal region. Wild-type and analog peptides were chemotactic for granulocytes and monocytes, irrespective of the salt concentrations. These new peptides may have therapeutic potential.Beta-defensins (BDs) are highly conserved small peptides produced by plants, invertebrates, and vertebrates that developed as part of the primordial immune protective mechanism (19). Four of these peptides, called human BD1 (hBD1; DEFB1), hBD2 (DEFB4), hBD3 (DEFB103A), and hBD4 (DEFB104), are mainly expressed by respiratory, gastrointestinal, and urogenital epithelial cells either constitutively (hBD1) or after induction by microorganisms or inflammatory factors (hBD2 to hBD4) (19). All four hBDs are cationic and 36 to 45 amino acids long and show similar folding and an invariable six-cysteine motif that gives rise to three disulfide bonds (2, 11, 12, 25, 26).Human beta-defensins 1 to 4 exert different bactericidal and antiviral activities against various pathogens (8, 15, 27). The antibacterial effects of hBD1 (9), hBD2 (33), and hBD4 (5) are attenuated by high NaCl concentrations, such as those in the airway surface fluid of patients with cystic fibrosis (CF) (21, 29). Human beta-defensin 3 can withstand NaCl concentrations as high as 150 mM, thanks to its peculiar structural characteristics and charge (10). In the field of viral diseases, hBD2 and -3 inhibit human immunodeficiency virus (HIV) type 1 (HIV-1) replication and virion infectivity (20, 31) and modulate HIV-1 coreceptor expression (20). Human herpes simplex virus (HSV) type 1 (HSV-1), HSV-2, and other viruses preincubated with alpha human neutrophil peptide 1 (hNP1) to hNP3 (6, 28) or theta (37) defensins lose their ability to infect target cells (28). As yet, there are no data on the effect of hBDs on HSV-1 and -2. In addition to direct antimicrobial activity, hBDs also exert chemotactic activity: hBD1, -2, and -3 are chemotactic for monocytes and dendritic and T cells. Human beta-defensin 3 is the only beta-defensin chemotactic for macrophages (4, 18, 19), whereas the chemotactic effect of hBDs on granulocytes has yet to be elucidated (4, 18).The two natural defensins hBD1 and hBD3 were chosen for use in the experiments described in this paper for the following reasons: hBD1 is constitutively expressed but its antibacterial activity is greatly impaired by NaCl, while hBD3 is insensitive to salt. Thus, we designed and synthesized hBD analogs that, in principle, would maintain the antibacterial and antiviral activities of hBD1 and possess a resistance capability in the presence of high NaCl concentrations, like hBD3 does. We then compared the antibacterial, chemotactic, and antiviral activities of the novel synthetic analogs with those of wild-type hBD1 and hBD3. Our data show that some of the synthetic analogs have higher antimicrobial activity than the wild type, also at high NaCl concentrations.     

Randomized double-blind comparison of serotonergic (Citalopram) versus noradrenergic (Reboxetine) reuptake inhibitors in outpatients with somatoform, DSM-IV-TR pain disorder.

OBJECTIVES: Whether the effect of tricyclic antidepressants on Pain Disorder arises from their noradrenergic or serotonergic actions or both remains unclear. We compared the selective serotonin reuptake inhibitor (SSRI) citalopram and the noradrenergic reuptake inhibitor (NARI) reboxetine in outpatients with Pain Disorder. We also distinguished the drugs' analgesic and antidepressant effects. METHODS: In this 8-week, randomized double-blind study, 35 patients with a DSM-IV-TR diagnosis of Pain Disorder were randomly assigned to receive either citalopram 40 mg/day (N=17 patients) or reboxetine 8 mg/day (N=18). The Present Pain Intensity (PPI) scale and the Total Pain Rating Index (tPRI) of the McGill Pain Questionnaire were used to measure the effect on pain symptoms. Changes in the Zung Self-Rating Depression Scale (Zung-D) scores were evaluated to monitor a possible antidepressant effect. For all patients who had at least one assessment, an intent-to-treat analysis was performed. RESULTS: No significant differences were found in the demographic variables or clinical characteristics of the two treatment groups. In the citalopram group, PPI and tPRI scores measured at baseline decreased after treatment (tPRI: 41.9 vs. 30.0, p=.004; PPI: 3.5 vs. 2.8, p=.045) whereas in the reboxetine group differences were not statistically significant (tPRI: 35.2 vs. 31.5; PPI: 3.7 vs. 3.1). The Zung-D showed no significant changes between baseline and endpoint assessment in either group. CONCLUSIONS: Our study suggests that the SSRI citalopram may have a moderate analgesic effect in patients with Pain Disorder, and that this analgesic activity appears to be not correlated to changes in depressive scores. If confirmed in a larger sample, this evidence suggests that patients who are intolerant or resistant to tricyclic antidepressants, may be treated with SSRIs.     

Cerebral Blood Flow in Migraine with Aura: A Transcranial Doppler Sonography Study

Transcranial Doppler sonography examination was performed on 44 patients with migraine with aura and 88 controls. All patients were investigated in headache-free periods and 10 of them also during a migraine attack. During the headache-free period a not significant increase of mean flow velocity in patients compared to controls was obtained. The pulsatility index (PI) mean values were also higher in patients than in controls and the differences were significant in the MCA (p < 0.05). No difference between right and left side was observed. During the attack the mean flow velocity (MFV) decreased in all arteries but the decrease was significant only in MCA and ACA (p < 0.05). The mean PI increased in all arteries but not significantly. These variations were observed both on the headache and contralateral side. Nevertheless, the MFV decrease in all arteries was observed in four patients only. In four patients the MFV decrease was found in the anterior arteries and the MFV increase in the posterior arteries, while in two patients the MFV increase was observed both in the anterior and posterior arteries. The correlation between the variations of MFV values during the attacks and the time interval from the onset of attacks showed that the PCA and BA flow velocities were increased in patients examined between 0.5 and 3 hours, while an increase in MCA and ACA flow velocities were observed only in patients examined after 1.5 hours.     

A human immunodeficiency virus type 1 pol gene-derived sequence (cPPT/CTS) increases the efficiency of transduction of human nondividing monocytes and T lymphocytes by lentiviral vectors

We have investigated the capacity of two human immunodeficiency virus type 1-derived lentivectors, differing in the presence of a 118-bp pol fragment containing the cPPT/CTS element, to transduce human normal primary cells of different hematopoietic lineages. Infection of resting monocytes with a high multiplicity of infection (MOI > 10) revealed that the lentivirus carrying the pol fragment (cPPT) is effective, transducing 75% of cells compared with 36% for the no-cPPT vector. Even at low MOIs (< or =1) the cPPT vector still shows a better transduction efficiency than the no-cPPT vector. Moreover, transduction does not require dendritic cell differentiation. In contrast, infection of nonactivated T lymphocytes showed that both vectors, tested at high MOIs, can transduce a small, although measurable, percentage of cells (up to 10%), which may correspond to G(1a) "activated" cells as detected by simultaneous staining of DNA and RNA, in our cultures in the presence of medium alone. Furthermore, we show that the sole addition of interleukin 2 or interleukin 15 represents a full proliferative signal under our conditions and permits high transduction efficiency (up to 30% with the cPPT vector and 15% with the no-cPPT vector). Still higher transduction of T lymphocytes can be achieved after stimulation with phytohemagglutinin and interleukin 2 (up to 78% with the cPPT vector vs. 42% with the no-cPPT vector). Finally, both viruses do not transduce either resting or proliferating tonsillar B lymphocytes.