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author:("colloidal, Giulia")
1.  Antioxidants inhibit SAA formation and pro-inflammatory cytokine release in a human cell model of alkaptonuria 
Rheumatology (Oxford, England)  2013;52(9):1667-1673.
Objective. Alkaptonuria (AKU) is an ultra-rare autosomal recessive disease that currently lacks an appropriate therapy. Recently we provided experimental evidence that AKU is a secondary serum amyloid A (SAA)-based amyloidosis. The aim of the present work was to evaluate the use of antioxidants to inhibit SAA amyloid and pro-inflammatory cytokine release in AKU.
Methods. We adopted a human chondrocytic cell AKU model to evaluate the anti-amyloid capacity of a set of antioxidants that had previously been shown to counteract ochronosis in a serum AKU model. Amyloid presence was evaluated by Congo red staining. Homogentisic acid-induced SAA production and pro-inflammatory cytokine release (overexpressed in AKU patients) were evaluated by ELISA and multiplex systems, respectively. Lipid peroxidation was evaluated by means of a fluorescence-based assay.
Results. Our AKU model allowed us to prove the efficacy of ascorbic acid combined with N-acetylcysteine, taurine, phytic acid and lipoic acid in significantly inhibiting SAA production, pro-inflammatory cytokine release and membrane lipid peroxidation.
Conclusion. All the tested antioxidant compounds were able to reduce the production of amyloid and may be the basis for establishing new therapies for AKU amyloidosis.
doi:10.1093/rheumatology/ket185
PMCID: PMC3741479  PMID: 23704321
ascorbic acid; N-acetylcysteine; taurine; phytic acid; lipoic acid; homogentisic acid; serum amyloid A; inflammation; chondrocyte; lipid peroxidation
2.  Effects of regenerative radioelectric asymmetric conveyer treatment on human normal and osteoarthritic chondrocytes exposed to IL-1β. A biochemical and morphological study 
Purpose
Osteoarthritis (OA) is a degenerative disease characterized by a progressive loss of articular cartilage extracellular matrix and is due to functional impairments occurring in chondrocytes. In previous works, we highlighted that Regenerative Tissue Optimization (TO-RGN) treatment with radioelectric asymmetric conveyer (REAC) technology influenced the gene expression profiles controlling stem cell differentiation and the pluripotency of human skin-derived fibroblasts in vitro. Since interleukin-1 beta signaling has been implicated in the induction and progression of this disease (through metalloproteinase-3 synthesis and nitric oxide production), we investigated whether REAC TO-RGN might influence the biochemical and morphological changes induced by interleukin-1 beta in normal and OA chondrocytes.
Methods
The induction of metalloproteinase-3 and proteoglycan synthesis was evaluated by a solid-phase enzyme-amplified sensitivity immunoassay, and nitric oxide production was evaluated with the Griess method. Ultrastructural features were observed by transmission electron microscopy.
Results
REAC TO-RGN treatment decreased nitric oxide and metalloproteinase-3 production in normal and OA chondrocytes, while inducing an increase in proteoglycan synthesis. OA chondrocytes were more affected by REAC TO-RGN treatment than were normal chondrocytes. Ultrastructural changes confirmed that REAC TO-RGN may counteract the negative effects of interleukin-1 beta incubation.
Conclusion
The results of this in vitro study suggest that REAC TO-RGN treatment may represent a new, promising approach for the management of OA.
doi:10.2147/CIA.S42229
PMCID: PMC3653677  PMID: 23682210
human chondrocytes ultrastructure; metalloproteinase; nitric oxide; proteoglycans; REAC TO-RGN treatment
3.  Polysorbate 80 and Helicobacter pylori: a microbiological and ultrastructural study 
BMC Microbiology  2012;12:217.
Background
The frequent occurrence of chemoresistant strains reduces the chances of eradication of H. pylori infection and prompted the investigation of non-antibiotic substances active against this organism. Some surfactants enhance the effectiveness of antibiotics for their permeabilizing properties towards bacteria. We examined the antimicrobial activity to H. pylori of the surfactant polysorbate 80, used alone and in association with amoxicillin, clarithromycin, metronidazole, levofloxacin and tetracycline. We also aimed to study the ultrastructural alterations caused upon H. pylori by polysorbate 80, alone and in combination with antibiotics. Twenty-two H. pylori strains were tested using the broth dilution method. After incubation, broth from each dilution was subcultured onto agar enriched with foetal bovine serum to determine the minimum bactericidal concentration (MBC). Synergistic effect of polysorbate 80 with antibiotics was investigated by the broth dilution and disc diffusion techniques. Ultrastructural alterations of organisms treated with polysorbate 80, alone and in association with antibiotics were analyzed by transmission electron microscopy.
Results
MBCs of polysorbate 80 ranged from 2.6 (1.1) μg/ml to 32 (0) μg/ml. Polysorbate 80 exerted a synergistic effect when associated with metronidazole and clarithromycin: polysorbate 80 and metronidazole MBCs decreased by ≥ 4 fold; clarithromycin MBCs for two resistant strains decreased by 20 and 1000 times. The principal alteration caused by polysorbate 80 consisted in the detachment of the outer membrane of bacteria.
Conclusions
The bactericidal activity of polysorbate 80 and the synergistic effect of the association with metronidazole and clarithromycin could be useful in the treatment of H. pylori infection.
doi:10.1186/1471-2180-12-217
PMCID: PMC3537525  PMID: 22998649
Bacteria; Antibiotics; Chemoresistances; Polysorbate 80; Transmission electron microscopy
4.  Genetic variation in SPAG16 regions encoding the WD40 repeats is not associated with reduced sperm motility and axonemal defects in a population of infertile males 
BMC Urology  2012;12:27.
Background
SPAG16 is a critical structural component of motile cilia and flagella. In the eukaryotic unicellular algae Chlamydomonas, loss of gene function causes flagellar paralysis and prevents assembly of the “9 + 2” axoneme central pair. In mice, we have previously shown that loss of Spag16 gene function causes male infertility and severe sperm motility defects. We have also reported that a heterozygous mutation of the human SPAG16 gene reduces stability of the sperm axonemal central apparatus.
Methods
In the present study, we analyzed DNA samples from 60 infertile male volunteers of Western European (Italian) origin, to search for novel SPAG16 gene mutations, and to determine whether increased prevalence of SPAG16 single nucleotide polymorphisms (SNPs) was associated with infertility phenotypes. Semen parameters were evaluated by light microscopy and sperm morphology was comprehensively analyzed by transmission electron microscopy (TEM).
Results
For gene analysis, sequences were generated covering exons encoding the conserved WD40 repeat region of the SPAG16 protein and the flanking splice junctions. No novel mutations were found, and the four SNPs in the assessed gene region were present at expected frequencies. The minor alleles were not associated with any assessed sperm parameter in the sample population.
Conclusions
Analysis of the SPAG16 regions encoding the conserved WD repeats revealed no evidence for association of mutations or genetic variation with sperm motility and ultrastructural sperm characteristics in a cohort of Italian infertile males.
doi:10.1186/1471-2490-12-27
PMCID: PMC3487941  PMID: 22963137
Sperm ultrastructure; Axoneme; Motile cilia; Male infertility; Central apparatus; Semen analysis
5.  The presence of bacteria species in semen and sperm quality 
Purpose
To verify the prevalence of semen bacterial contamination and whether the contamination could decrease sperm quality.
Methods
Spermiogram, semen culture, and sperm transmission electron microscopy (TEM) analysis were performed. TEM data were elaborated using a mathematical formula that calculates a fertility index (FI)—able to define patients as fertile or infertile—and the percentage of sperm apoptosis, immaturity and necrosis. We aligned the amino acid sequence of beta-tubulin with protein of the most frequent species isolated from semen.
Results
Patients were divided according to the contaminating species; in each group, we observed fertile individuals, in whom the semen quality was similar to that of controls and infertile men whose sperm quality was significantly decreased, in terms of motility, FI, apoptosis and necrosis. Partial homology between β-tubulin and bacterial proteins was observed.
Conclusion
Sperm bacterial contamination is quite frequent and could contribute to the deterioration of the sperm quality of infertile men.
doi:10.1007/s10815-008-9283-5
PMCID: PMC2649329  PMID: 19089609
Bacteria; Semen quality; Spermatozoa; TEM
6.  18, X, Y aneuploidies and transmission electron microscopy studies in spermatozoa from five carriers of different reciprocal translocations 
Asian Journal of Andrology  2009;11(3):325-332.
We analysed ejaculated spermatozoa from five infertile men with different balanced reciprocal translocations to contribute to the study of meiotic segregation of chromosomes 18, X and Y and also to evaluate sperm morphology by transmission electron microscopy (TEM) analysis. Conventional lymphocyte karyotype analyses highlighted different reciprocal balanced translocations: t(12;13), t(4;9), t(X;8), t(8;10) and t(3;16). Semen analysis was performed by light and TEM. Fluorescence in situ hybridization was performed directly on sperm nuclei using centromeric probes for chromosomes 18, X and Y. The carriers of the balanced reciprocal translocations considered in the present study showed a very similar pattern of sperm pathologies: diffused presence of apoptosis and immaturity. All patients showed meiotic segregation derangements, highlighted by the presence of sperm diploidies and sex chromosome disomies particularly related to the failure of the first meiotic division. However, an increased incidence of chromosome 18 aneuploidy was detected in spermatozoa from t(X;8) and t(8;10) carriers. We have also reported values from sex chromosomes such as t(X;8), although the X chromosome was involved in translocation. Since patients with reciprocal translocations and spermatogenetic impairment are candidates for intracytoplasmic sperm injection cycles, the study of sperm parameters, and particularly of the level of aneuploidy rates, would provide better information for couples at risk and would contribute to the data in the literature for a better understanding of the effects of chromosomal rearrangement on the whole meiotic process and, in particular, on chromosomes not involved in translocation.
doi:10.1038/aja.2008.31
PMCID: PMC3735288  PMID: 19349951
electron microscopy; fluorescence in situ hybridization; altered karyotype; spermatozoa; reciprocal translocation

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