Azoospermia: Is it worth waiting for the confirmation of the semen abnormality to start an infertility assessment?

Azoospermia is found in about 1% of men in the general population and in about 10%-15% of infertile men. Upon discovery of semen analysis abnormality, another test must be performed after an interval of 3 months before any other infertility work-up. This research aimed at evaluating the benefit of waiting for the control test. This retrospective monocentric descriptive study was carried out in the fertility center of the University Hospital of Saint Etienne. All consecutive azoospermic patients diagnosed between January, 2012 and December, 2019 were included. For each patient, two consecutive semen analyses performed 3 months apart were studied. The main focas was on patients whose second semen analysis would have modified the infertility work-up. Amongst the 172 cases under study, the second semen analysis revealed the presence of sperm for three men. Only one of these 3 modified semen analyses was normal. In the observed azoospermic population, sperm was found on the second test in 1.7%. An infertility assessment is necessary after the discovery of azoospermia in the first semen analysis in 99.5%. These results suggest that it is useless to wait three stressful months before starting an infertility assessment for azoospermic population.

Impact of Nanoparticles on Male Fertility: What Do We Really Know? A Systematic Review.

The real impact of nanoparticles on male fertility is evaluated after a careful analysis of the available literature. The first part reviews animal models to understand the testicular biodistribution and biopersistence of nanoparticles, while the second part evaluates their in vitro and in vivo biotoxicity. Our main findings suggest that nanoparticles are generally able to reach the testicle in small quantities where they persist for several months, regardless of the route of exposure. However, there is not enough evidence that they can cross the blood-testis barrier. Of note, the majority of nanoparticles have low direct toxicity to the testis, but there are indications that some might act as endocrine disruptors. Overall, the impact on spermatogenesis in adults is generally weak and reversible, but exceptions exist and merit increased attention. Finally, we comment on several methodological or analytical biases which have led some studies to exaggerate the reprotoxicity of nanoparticles. In the future, rigorous clinical studies in tandem with mechanistic studies are needed to elucidate the real risk posed by nanoparticles on male fertility.

Quantitative Flow Cytometric Evaluation of Oxidative Stress and Mitochondrial Impairment in RAW 264.7 Macrophages after Exposure to Pristine, Acid Functionalized, or Annealed Carbon Nanotubes.

Conventional nanotoxicological assays are subjected to various interferences with nanoparticles and especially carbon nanotubes. A multiparametric flow cytometry (FCM) methodology was developed here as an alternative to quantify oxidative stress, mitochondrial impairment, and later cytotoxic and genotoxic events. The experiments were conducted on RAW264.7 macrophages, exposed for 90 min or 24 h-exposure with three types of multiwalled carbon nanotubes (MWCNTs): pristine (Nanocyl™ CNT), acid functionalized (CNTf), or annealed treatment (CNTa). An original combination of reactive oxygen species (ROS) probes allowed the simultaneous quantifications of broad-spectrum ROS, superoxide anion (O2•-), and hydroxyl radical (•OH). All MWCNTs types induced a slight increase of broad ROS levels regardless of earlier antioxidant catalase activity. CNTf strongly stimulated the O2•- production. The •OH production was downregulated for all MWCNTs due to their scavenging capacity. The latter was quantified in a cell-free system by electron paramagnetic resonance spectroscopy (EPR). Further FCM-based assessment revealed early biological damages with a mitochondrial membrane potential collapse, followed by late cytotoxicity with chromatin decondensation. The combined evaluation by FCM analysis and cell-free techniques led to a better understanding of the impacts of MWCNTs surface treatments on the oxidative stress and related biological response.

Ex vivo detection and quantification of gold nanoparticles in human seminal and follicular fluids.

Increasing consumption of engineered nanoparticles and occupational exposure to novel, ultrafine airborne particles during the last decades has coincided with deterioration of sperm parameters and delayed fecundity. In order to prevent possible adverse health effects and ensure a sustainable growth for the nanoparticle industry, the ability to investigate the nanosized, mineralogical load of human reproductive systems is becoming a real clinical need. Toward this goal, the current study proposes two methods for the detection and quantification of engineered nanoparticles in human follicular and seminal fluid, developed with the use of well-defined 60 nm Au particles. Despite the complexity of these biological fluids, simple physical and chemical treatments allow for the precise quantification of more than 50 and 70% wt of the spiked Au nanoparticles at low µg ml-1 levels in follicular and seminal fluids, respectively. The use of electron microscopy for the detailed observation of the detected analytes is also enabled. The proposed method is applied on a small patient cohort in order to demonstrate its clinical applicability by exploring the differences in the metal and particulate content between patients with normal and low sperm count.

Affinity of Indocyanine Green in the Detection of Colorectal Peritoneal Carcinomatosis.

Indocyanine green (ICG) is increasingly being used in digestive oncology. In colorectal cancer, ICG can be used to detect lymph node metastasis and hepatic metastasis on the surface of the liver. In peritoneal carcinomatosis, it was previously suspected that the diffusion of ICG in the tumor mass was due to the enhanced permeability and retention effect; however, this phenomenon has not been clearly demonstrated. Using bevacizumab, an antibody directed against vascular endothelial growth factor that consequently inhibits neoangiogenesis, we sought to confirm the mode of ICG diffusion. We compared the fluorescence of peritoneal carcinomatosis nodules from patients who had previously received bevacizumab during their oncologic treatment with those who did not receive this therapy. The sensitivity of the carcinomatosis nodule fluorescence was higher in the patients who did not receive bevacizumab compared with those who received the drug (76.3% and 65.0%, respectively). The rate of false-negative results was higher in the bevacizumab group than in the group that did not receive the drug (53.8% and 42.9%, respectively). Using bevacizumab, we demonstrate that the enhanced permeability and retention effect causes ICG accumulation in peritoneal carcinomatosis resulting from colorectal cancer.

Pulsatile gonadotropin-releasing hormone therapy in persistent amenorrheic weight-recovered anorexia nervosa patients.

OBJECTIVE: To compare hormonal and clinical responses to GnRH pulsatile treatment in weight-recovered anorexia nervosa patients (Rec-AN) with persistent functional hypothalamic amenorrhea (HA) vs. in patients with secondary and primary HA. DESIGN: Retrospective, observational, ambulatory study. SETTING: University hospital. PATIENT(S): Forty-one women: 19 Rec-AN (body mass index >18.5 kg/m2 without menses recovery), 15 secondary HA without any eating disorders patients (SHA), and 7 primary HA patients (PHA). INTERVENTION(S): Gonadotropin-releasing hormone pulsatile therapy. MAIN OUTCOME MEASURE(S): Baseline E2, LH, and P plasma levels and their changes during induction cycles; ovulation, follicular recruitment, and pregnancies. RESULTS: The Rec-AN group displayed higher basal E2 and LH plasma levels after GnRH injection compared with SHA and PHA. Higher E2 and LH levels were observed during induction cycles in Rec-AN compared with SHA and PHA. Follicular recruitment was higher in Rec-AN. The ovulation rate was higher in Rec-AN compared with PHA but similar to SHA. CONCLUSION(S): This study showed increased gonadal status and higher E2 response to pulsatile GnRH therapy in persistent amenorrheic weight-recovered AN compared with HA from other causes. It suggests that their individual set-point of body weight allowing a fully functional gonadal axis is not reached yet. Specific factors of gonadal inertia in Rec-AN still remain unclear.

Detection and analysis of nanoparticles in patients: A critical review of the status quo of clinical nanotoxicology.

On the cusp of massive commercialization of nanotechnology-enhanced products and services, the physical and chemical analysis of nanoparticles in human specimens merits immediate attention from the research community as a prerequisite for a confident clinical interpretation of their occurrence in the human organism. In this review, we describe the caveats in current practices of extracting and isolating nanoparticles from clinical samples and show that they do not help truly define the clinical significance of detected exogenous nano-sized objects. Finally, we suggest a systematic way of tackling these demanding scientific tasks. More specifically, a precise and true qualitative evaluation of nanoparticles in human biological samples is still hindered by various technical reasons. Such a procedure is more refined when the nature of the pollutants is known, like in the case of nano-sized wear debris originating from biomedical prostheses. Nevertheless, nearly all available analytical methods provide unknown quantitative accuracy and qualitative precision due to the challenging physical and chemical nature of nanoparticles. Without trustworthy information to describe the nanoparticulate load of clinical samples, it is impossible to accurately assess its pathological impact on isolated cases or allow for relevant epidemiological surveys on large populations. Therefore, we suggest that the many and various specimens stored in hospitals be used for the refinement of methods of exhaustive quantitative and qualitative characterization of prominent nanoparticles in complex human milieu.

Testicular biodistribution of silica-gold nanoparticles after intramuscular injection in mice.

With the continuing development of nanomaterials, the assessment of their potential impact on human health, and especially human reproductive toxicity, is a major issue. The testicular biodistribution of nanoparticles remains poorly studied. This study investigated whether gold-silica nanoparticles could be detected in mouse testes after intramuscular injection, with a particular focus on their ability to cross the blood-testis barrier. To that purpose, well-characterized 70-nm gold core-silica shell nanoparticles were used to ensure sensitive detection using high-resolution techniques. Testes were collected at different time points corresponding to spermatogenesis stages in mice. Transmission electron microscopy and confocal microscopy were used for nanoparticle detection, and nanoparticle quantification was performed by atomic emission spectroscopy. All these techniques showed that no particles were able to reach the testes. Results accorded with the normal histological appearance of testes even at 45 days post sacrifice. High-resolution techniques did not detect 70-nm silica-gold nanoparticles in mouse testes after intramuscular injection. These results are reassuring about the safety of nanoparticles with regard to male human reproduction, especially in the context of nanomedicine.

Metals distribution in colorectal biopsies: New insight on the elemental fingerprint of tumour tissue.

BACKGROUND: Some studies have linked colorectal cancer to metal exposure. AIMS: Our objective was to evaluate the element distribution in colorectal adenocarcinoma biopsies, adjacent non-tumour tissues, and healthy controls. METHODS: The study is a case-control study which compared the element distribution in colon biopsies from two groups of patients: with colorectal cancer (2 types of samples: colorectal cancer biopsies and adjacent non-tumour tissues) and healthy controls. Fifteen metal concentrations (Aluminium, Boron, Cadmium, Chromium, Copper, Iron, Magnesium, Manganese, Nickel, Lead, Selenium, Silicon, Titanium, Vanadium, and Zinc) were quantified by using inductively coupled plasma atomic emission spectrometry. RESULTS: 104 patients were included: 76 in the colorectal cancer group, 28 in the healthy control group. Among the 15 elements analyzed, only boron, chromium, zinc, silicon and magnesium were found at clearly detectable concentrations. Colorectal tumour biopsies had significantly higher concentrations of magnesium as compared to adjacent non-tumour or healthy tissues. Zinc concentration followed the same trend but differences were not statistically significant. In addition, concentration of silicon was higher in colorectal cancer tissue than in healthy non-cancer tissue, while chromium was mostly found in adjacent non-tumour tissue. CONCLUSION: Magnesium, chromium, zinc and silicon were found in noteworthy concentrations in colorectal tumour. Their potential role in colorectal carcinogenesis should be explored.

Quantification of nanoparticle endocytosis based on double fluorescent pH-sensitive nanoparticles.

Amorphous silica is a particularly interesting material because of its inertness and chemical stability. Silica nanoparticles have been recently developed for biomedical purposes but their innocuousness must be carefully investigated before clinical use. The relationship between nanoparticles physicochemical features, their uptake by cells and their biological activity represents a crucial issue, especially for the development of nanomedicine. This work aimed at adapting a method for the quantification of nanoparticle endocytosis based on pH-sensitive and double fluorescent particles. For that purpose, silica nanoparticles containing two fluorophores: FITC and pHrodo(TM) were developed, their respective fluorescence emission depends on the external pH. Indeed, FITC emits a green fluorescence at physiological pH and pHrodo(TM) emits a red fluorescence which intensity increased with acidification. Therefore, nanoparticles remained outside the cells could be clearly distinguished from nanoparticles uptaken by cells as these latter could be spotted inside cellular acidic compartments (such as phagolysosomes, micropinosomes…). Using this model, the endocytosis of 60 nm nanoparticles incubated with the RAW 264.7 macrophages was quantified using time-lapse microscopy and compared to that of 130 nm submicronic particles. The amount of internalized particles was also evaluated by fluorimetry. The biological impact of the particles was also investigated in terms of cytotoxicity, pro-inflammatory response and oxidative stress. Results clearly demonstrated that nanoparticles were more uptaken and more reactive than submicronic particles. Moreover, we validated a method of endocytosis quantification.

Long-term in vivo clearance of gadolinium-based AGuIX nanoparticles and their biocompatibility after systemic injection.

We previously reported the synthesis of gadolinium-based nanoparticles (NPs) denoted AGuIX (activation and guiding of irradiation by X-ray) NPs and demonstrated their potential as an MRI contrast agent and their efficacy as radiosensitizing particles during X-ray cancer treatment. Here we focus on the elimination kinetics of AGuIX NPs from the subcellular to whole-organ scale using original and complementary methods such as laser-induced breakdown spectroscopy (LIBS), intravital two-photon microscopy, inductively coupled plasma optical emission spectrometry (ICP-OES), transmission electron microscopy (TEM), and electrospray ionization mass spectrometry (ESI-MS). This combination of techniques allows the exact mechanism of AGuIX NPs elimination to be elucidated, including their retention in proximal tubules and their excretion as degraded or native NPs. Finally, we demonstrated that systemic AGuIX NP administration induced moderate and transient effects on renal function. These results provide useful and promising preclinical information concerning the safety of theranostic AGuIX NPs.