Carbon dots, a powerful non-toxic support for bioimaging by fluorescence nanoscopy and eradication of bacteria by photothermia.

Carbon Dots (CDs) are innovative materials which have potential applications in many fields, including nanomedicine, energy and catalysis. Here CDs were produced by the alkali-assisted ultrasonic route and characterized by several techniques to determine their composition and properties. Fluorescence nanoscopy using single-molecule localization microscopy shows that they have very good photophysical properties and a remarkable blinking behaviour at 405 nm. Moreover, these CDs are a safe material, non-toxic towards different cell lines (cancer and non-cancer cells) even at very high concentration, reflecting an excellent biocompatibility. Photothermia, i.e. their heating capacity under laser irradiation, was evaluated at two wavelengths and at several power densities. The resulting temperature increment was high (5 < ΔT < 45 °C) and appropriate for biomedical applications. Bioimaging and photothermia were then performed on E. coli, a Gram(-) bacterium, incubated with CDs. Remarkably, by photothermia at 680 nm (0.3, 1 and 1.9 W cm-2) or 808 nm (1.9 W cm-2), CDs are able to eradicate bacteria in their exponential and stationary phases. Images obtained by 3D super-resolution microscopy clearly show the different CD distributions in surviving bacteria after mild photothermal treatment. These results confirm that CDs are multifunctional materials with a wide range of biomedical applications.

TRAIL-NP hybrids for cancer therapy: a review.

Cancer is a worldwide health problem. It is now considered as a leading cause of morbidity and mortality in developed countries. In the last few decades, considerable progress has been made in anti-cancer therapies, allowing the cure of patients suffering from this disease, or at least helping to prolong their lives. Several cancers, such as those of the lung and pancreas, are still devastating in the absence of therapeutic options. In the early 90s, TRAIL (Tumor Necrosis Factor-related apoptosis-inducing ligand), a cytokine belonging to the TNF superfamily, attracted major interest in oncology owing to its selective anti-tumor properties. Clinical trials using soluble TRAIL or antibodies targeting the two main agonist receptors (TRAIL-R1 and TRAIL-R2) have, however, failed to demonstrate their efficacy in the clinic. TRAIL is expressed on the surface of natural killer or CD8+ T activated cells and contributes to tumor surveillance. Nanoparticles functionalized with TRAIL mimic membrane-TRAIL and exhibit stronger antitumoral properties than soluble TRAIL or TRAIL receptor agonist antibodies. This review provides an update on the association and the use of nanoparticles associated with TRAIL for cancer therapy.

Stemness of spermatogonial stem cells encapsulated in alginate hydrogel during cryopreservation.

This study investigated the effect of spermatogonial stem cell encapsulated in alginate hydrogel during cryopreservation, as cells were protected against damage during cryopreservation within the hydrogel. Spermatogonial stem cells were isolated from the testes of Balb/c mice pups (6 days old), purified in laminin-coated dishes and CD90.1 microbeads, encapsulated in alginate hydrogel and then cryopreserved. After thawing, cell viability and Spermatogonial stem cell (SSC) colony diameter were evaluated. After RNA was isolated and cDNA was synthesised, the expression of stemness genes was considered using RT real-time PCR. Finally, spermatogonial stem cells labelled with BrdU were transplanted to busulfan azoospermic mouse models. Lin28a and Sall4 genes were significantly upregulated after cryopreservation in alginate hydrogel. However, cell viability was significantly decreased. The diameter of colonies consisting of spermatogonial stem cells freeze-thawed in alginate microbeads showed no significant difference with fresh spermatogonial stem cells and the control group. The injection of freeze-thawed spermatogonial stem cells encapsulated in alginate hydrogel resulted in spermatogenesis recovery. Alginate mimics the extracellular matrices (ECM) for spermatogonial stem cells; therefore, it can support stemness potential during the cell cryopreservation process and restart spermatogenesis after transplantation.

Maghemite nanoparticles coated with human serum albumin: combining targeting by the iron-acquisition pathway and potential in photothermal therapies.

Human serum albumin (HSA), the most abundant plasma protein in human blood, is a natural transport vehicle with multiple ligand binding sites. It, therefore, constitutes an attractive candidate for drug delivery. Targeting may occur via the most known interaction of the protein with the neonatal Fc receptor (FcRn). Here, we investigate another HSA delivery path, involving the transferrin receptor, and we elaborate a maghemite-HSA nanohybrid, opening up new opportunities for medical applications. Fluorescence spectrophotometric titration and size-exclusion chromatography were used to substantiate, in cell-free assays, an interaction between HSA and the transferrin receptor R1. This occurs with a dissociation constant, KD of 6.7 nM. This interaction was confirmed in HeLa cell culture where, by confocal microscopy, rhodamine-labeled HSA is shown to be internalized. HSA was then covalently conjugated onto maghemite nanoparticles (NPs) to give a NP-HSA nanohybrid. The therapeutic potential of this hybrid was demonstrated through its heating capacity in magnetic hyperthermia (MH) and near-infrared (NIR) photothermia (PT). In particular, the Specific Absorption Rate (SAR) in the PT Therapy (PTT) mode, using a 808 nm NIR-LASER (1 W cm-2) and at iron concentration as low as 2.5 mM, was found to be very high, equal to 1870 W g-1 with a temperature increment of 9.2 °C. The nanohybrids incubated with HeLa cells were mainly localized at the cell surface. When the PTT mode was applied under the same conditions as in vitro, mortality was higher in HeLa cells than in fibroblasts (non-malignant cells). Cytotoxicity was checked in both cell lines without the PTT mode; the nanohybrids do not seem to affect cell viability. These results make the nanohybrids very promising agents for NIR-PT and for targeting in cancer therapy, since non-malignant cells were not damaged.

Morphometric and stereological assessment of the effects of titanium dioxide nanoparticles on the mouse testicular tissue.

OBJECTIVE: To evaluate the effects of titanium dioxide nanoparticles (TNPs) on morphometric and stereological parameters of mouse testis. BACKGROUND: TNPs are increasingly used in sunscreens, biosensors, food additives, pigments, rubber manufacture, and electronic materials. However, the potential toxicity of these nanoparticles is not well understood. METHODS: Experimental Groups (TNP-1, TNP-2 and TNP-3) received one of the following treatments daily for 35 days: 75, 150 and 300 mg/kg TNPs respectively. Right testis from each animal was fixed in bouin's solution for measurement of total volume of testis, total volume of seminiferous tubules, total volume of interstitial tissue and total number of Leydig cells by stereological methods. Seminiferous tubule diameter and seminiferous epithelium height were assessed by morphometrical method. The left testicles were homogenized for measurement of testosterone concentration. RESULTS: There was a significant decrease in the weight of testicles in TNP-3 groups. The stereological and morphometrical parameters were significantly changed in TNP-2 and TNP-3 groups. TNP-2 and TNP-3 groups also showed a significant decrease in testosterone concentrations (p < 0.05). CONCLUSION: This study had demonstrated that TNPs could change stereological and morphometrical parameters of the seminiferous tubules and reduce the number of Leydig cells and testosterone concentration (Tab. 3, Fig. 3, Ref. 35).

Melatonin protects testes against lithium-pilocarpine-induced temporal lobe epilepsy in rats: a time course study.

Male dysfunction is common in patients with temporal lobe epilepsy (TLE). We evaluated whether melatonin, as a supplement, can play a positive role in reducing the epileptogenesis imposing abnormalities of spermatozoa and testes in epileptic rats. Status epilepticus was induced based on the TLE lithium-pilocarpine model. Two patterns of melatonin were administered to the epileptic animals along the mean durations of latent (14 days) and chronic (60 days) phases. Sperm parameters, different antioxidant enzyme levels, germ cell apoptosis, body and relative sex organ weights were evaluated in all groups 60 days following SE induction. Chronic TLE caused a significant reduction in sperm parameters. In the testis, the reduced level of antioxidant enzymes was accompanied by a significant increase in malondialdehyde concentration. The presence of oxidant condition in the testes of epileptic animals caused expanded apoptosis in the germ cell layer. Moreover, the amount of weight gain in epileptic animals was more prominent. Melatonin administration was able to improve sperm motility by increasing the total antioxidant level. There was also a significant reduction in the spermatogenic cell line apoptosis and the extra weight gain of melatonin-treated animals. Melatonin supplementation might be considered as an acceptable cotreatment in epileptic patients.

Design and synthesis of 3-isoxazolidone derivatives as new Chlamydia trachomatis inhibitors.

Chlamydia trachomatis (Ct) is a bacterial human pathogen responsible for the development of trachoma, the worldwide infection leading to blindness, and is also a major cause of sexually transmitted diseases. As iron is an essential metabolite for this bacterium, iron depletion presents a promising strategy to limit Ct proliferation. The aim of this study is to synthesize 3-isoxazolidone derivatives bearing known chelating moieties in an attempt to develop new bactericidal anti-Chlamydiaceae molecules. We have investigated the paths by which these new compounds affect Ct serovar L2 development in HeLa cells, in the presence or absence of exogenously added iron. The iron-chelating properties of these molecules were also determined. Our data reveal important bactericidal effects which are distinguishable from those due to iron chelation.

Combination of running exercise and high dose of anabolic androgenic steroid, nandrolone decanoate, increases protamine deficiency and DNA damage in rat spermatozoa.

High doses of anabolic-androgenic steroids (AAS) are used by some athletes to increase muscle mass, that is often associated with male infertility. The aim of this study was to investigate the possible cause/s of male infertility using a rat model by analysing sperm quality, including its protamine content and DNA integrity, as well as pregnancy rate. Five groups of male Wistar rats were treated for 10 weeks as follows: nandrolone decanoate (10 mg kg(-1) per week) (ND); running exercise (50 min per day, 5 days a week) (EX); Combination of ND and exercise (ND-EX); nandrolone decanoate solvent (Sham); and control without any injection or exercise (CO). Deterioration in sperm quantity was observed in all test groups (P ≤ 0.01). The frequency of fertile rats was decreased in the ND-EX and ND groups (P ≤ 0.05). Chromomycin-A3 staining showed a protamine deficiency in the epididymal spermatozoa in the ND-EX rats (P ≤ 0.05). Chromatin analysis indicated an abnormal maturation of the sperm nuclei in all test groups compared with the controls (P ≤ 0.05). TUNEL analyses showed a highly significant increase in apoptosis in the EX, ND, and ND-EX groups (P ≤ 0.01). Our data show that a combination of exercise and high doses of nandrolone decanoate negatively influences the DNA integrity and protamine content resulting in lower sperm quality and reduced pregnancy rate.

Potential use of melatonin supplementation to protect vitrified testicular grafts from hypoxic-ischaemic damage.

This study aimed to assess the morphological changes in neonate vitrified testicular grafts after host treatment with melatonin. Neonate vitrified testes, candidates for transplantation to treated and nontreated groups receiving melatonin, were thawed in media containing or not containing 100 µm melatonin. Following transplantation, melatonin (20 mg kg(-1) per day) and saline were given to the treated and nontreated groups for up to 9 weeks. The testicular status was assayed by terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP)-biotin nick-end labelling TUNEL, semi-thin section and ultra-structure studies. Chi-squared test was used to compare categorical variables between the groups. Overall, the degenerative and apoptosis changes in the vitrified testis parenchyma were not significant. However, atrophic seminiferous cords and jumbled appearance of the interstitial space were more often observed in the nontreated group than in the treated ones. Semi-thin sections showed that the germinal epithelium was taken in a normal arrangement on the testicular grafts of both groups. Nevertheless, ultrastructural analysis revealed that the characteristics of interstitial space cells, basement membrane BM and epithelial cells of seminiferous tubules in the treated group were better than those in the nontreated group. The study revealed a beneficial effect of melatonin on vitrified neonatal testes and after that, on restoring testicular grafts.

Assessment of morphological and functional changes in neonate vitrified testis grafts after host treatment with melatonin.

This study was conducted to assess the effect of melatonin on the structure of testis and spermatogenesis dynamics in neonate vitrified testis grafts. Neonate vitrified testes, candidates for transplantation heterotopically to experiment or control groups, were warmed in thawing media which had or did not have a supplement of 100 µM melatonin, respectively. Following transplantation, melatonin (20 mg/ /kg/day) or saline solution was intraperitoneally injected into the treated and the non-treated groups, respectively. The initiating spermatogenesis, spermatogonia survival, and structure of tissue in the testis graft were examined. Cell apoptosis (TUNEL assay) and proliferation (Brdu assay) in germ cells were determined. Histological studies revealed the dynamic of the spermatogenesis process in the vitrified testis graft. However, dilation of the lumen accompanied by a disorganised epithelium in the non-treated group was higher than in the treated group. Furthermore, the proportion of apoptotic germ cells together with a reduced proportion of proliferated germ cells was higher in the non-treated group than in the treated group. Overall, the number of seminiferous tubules in the testes grafts of both groups remained steady. However, the non-treated testes grafts contained more damaged seminiferous tubules than the treated ones. The thickness of the seminiferous tubules was greater in the melatonin treated group than in the non-treated group. In fact, the thickness of germinal epithelium was significantly higher in the treated group than in the non-treated group. The study may show a positive effect from melatonin resulting in more grafts restoring puberty. Furthermore, the associated increase in the healthy number of seminiferous tubules suggests that melatonin may have a preventative ischaemia/antioxidant role and in fact may be useful to initiate the spermatogenesis process.

Follicular dynamics in neonate vitrified ovarian grafts after host treatment with melatonin.

This study evaluates the effect of melatonin on follicular dynamics in neonate vitrified ovarian grafts. Vitrified ovaries from neonate F1 hybrid mice, candidates for transplantation to treated or non-treated groups, were thawed under standard conditions with or without the addition of 100 µM melatonin, respectively. Following transplantation, melatonin (20 mg/kg/day) or saline solution was injected i.p. into the treated and the non-treated groups, respectively. Vaginal cytology to monitor estrogenic activity together with follicle survival and development in the ovary grafts was examined. The results showed that the sites of transplantation became obvious within the oestrous phase. Histological analysis showed that there was a dynamic of the ovogenesis process in the vitrified ovary grafts. However, in the ovary graft the empty cavity together with jumbled oocyte-granulosa complex in the non-treated group was higher than in the melatonin treated group. Overall, the number of primary follicles in the ovary grafts of both groups remained constant throughout the oestrous period. However, the treated ovary grafts contained more secondary follicles throughout the oestrous period in comparison to non-treated ovary grafts. The antral follicle rates were more marked in the melatonin treated group than in the non-treated group during the oestrous period, whereas at other days of the cycle no significant difference was observed. The fresh corpora lutea rates were significantly higher in the treatment group than in the non-treatment group. The study showed that there is a positive effect with melatonin resulting in more grafts restoring puberty. Furthermore, the associated increase in healthy follicles suggests that melatonin has a preventative ischaemia/antioxidant action and may be useful to follicles.

The effects of swimming exercise and supraphysiological doses of nandrolone decanoate on the testis in adult male rats: a transmission electron microscope study.

Anabolic-androgenic steroids (AAS) are used in high doses by athletes to improve athletic ability, physical appearance, and muscle mass. Unfortunately, the abuse of these agents has significantly increased. It has been established that exercise and high doses of AAS may influence the hypothalamic-pituitary gonadal (H-P-G) axis, which can in turn affect the ultrastructure of the testes. However, the effect of the combination of exercise and high doses of AAS on the ultrastructure of the testes is not known. This study was undertaken in order to examine the combination effects of swimming exercise and supraphysiological doses of nandrolone decanoate on the ultrastructural changes in rat testes. Five groups of male Wistar strain albino rats were treated as follows for 8 weeks: solvent of nandrolone decanoate (peanut oil) as a vehicle (sham); nandrolone decanoate (ND) (10 mg/kg/week) - ND; exercise (1 h/day, 5 days a week) - exercise; ND (10 mg/kg/week) and exercise (1 h/day, 5 days a week) - ND-EX; and sedentary control without any injection or exercise - control. Ultrastructural changes in the rat testes were characterised by transmission electron microscopy. The number and size of Leydig cells were considerably decreased in the interstitial space in the experimental rats. The increased thickness and irregular wavy multilaminar appearance of basement membrane in the treated animals, especially in the ND-EX group, are associated with well developed myoid cells. Cytoplasm vacuolisation, vesicular-like crista of the mitochondria, numerous lipid droplets, and lysosome and phagolysosome in Sertoli cells were significantly observed in the experimental groups. Several apoptotic germ cells were considerably observed in the experimental rats (p ≤ 0.05). Exercise training seems to increase the extent of ultrastructural changes caused by supraphysiological doses of ND in rats, which in turn may affect fertility.