Microfluidic single sperm analysis

Microfluidic technology has been occasionally used for in vitro analysis and separation of cells. The small dimensions of microfluidic chips are very suitable to study cells on the single cell level rather than in whole populations. Also sperm cells have been studied and manipulated using microfluidic setups. Studying these cells on the single cell level is relevant for applications in assisted reproduction technologies and for sperm refinement in the livestock industries. \ud In this thesis, two different setups are proposed to study sperm cells individually. The first setup was designed to entrap a sperm cell on a fixed position. It consists of a microfluidic chip, which consists of two main channels that are interconnected by tiny side channels. Upon a pressure difference in between these main channels, the sperm cells were trapped since their head is too large to fit through. Now, several sperm characteristics including the integrity of the plasma and acrosomal membranes were studied on the single cell level using fluorescent staining. This system was also used to electrically study the swimming behaviour of individual trapped sperm cells using integrated microelectrodes. Recorded changes in impedance provided information upon the sperm swimming behaviour. The ability to study individual sperm cells in a non-destructing way has potential for improving sperm selection in intracytoplasmic sperm injection treatments. \ud The second setup consists of a single channel in which sperm cells are flown through a pair of microelectrodes. These electrodes were used to electrically analyse each passing sperm cell. The recorded electrical information can provide information upon the integrity of the plasma membrane and the cell morphology. In this thesis, the presence of a morphological sperm anomaly (cytoplasmic droplet) was studied and successfully identified. Furthermore, a label-free cell sorting system was developed in which sperm cells can be sorted using dielectrophoresis based on impedance data. Potentially, this sorting system is able to sort sperm cells in a non-destructing way based on cell viability and morphology. This system is very interesting for the livestock industry, in which removal of abnormal sperm cells from semen can improve semen quality and the outcome of artificial insemination treatments

Microfluidic device for selection of semen

The invention provides a system (1) for performing sperm analysis and selection based on sperm cell morphology of sperm cells (6) in a fluid (5), the system (1) comprising: (i) a fluid flow channel (2) for transport of said fluid (5), the fluid flow channel (2) comprising an inlet (10) an analyzing zone (40) configured downstream from said inlet (10) and comprising a first pair of electrodes (41) comprising a first intra-electrode distance (dl), a sorting zone (50) configured downstream from said analyzing zone (40) and comprising a sorting device (51), and outlets (80, 90, . . . ) configured downstream from said sorting zone (50); (ii) an electric source (140) configured to provide an electric signal to the first pair of electrodes (41); (iii) a measuring device (150) functionally coupled to the first pair of electrodes (41) and configured to measure a first impedance as a function of time of the fluid (5) between the first pair of electrodes, and to provide time-dependent impedance data; wherein the sorting device (51) is configured to sort sperm cells (6) by directing the sperm cell (6) in the sorting zone (50) to one of the outlets (80, 90, . . . ) based on a comparison in a comparison stage of the time dependent impedance data with predefined reference data

Spermometer: electrical characterization of single boar sperm motility

Objective:\ud To study single sperm boar motility using electrical impedance measurements in a microfluidic system. \ud \ud Design:\ud Comparison of the optical data and electrical impedance data. \ud \ud Setting:\ud Research laboratory at a university. \ud \ud Animal(s):\ud Boar semen sample were used. \ud \ud Intervention(s):\ud A microfluidic system is developed that is able to spatially confine single boar sperm cells and allows noninvasive analysis of their motility on the single cell level. Using this system, the single sperm motility was affected by changing the temperature or adding chemical stimuli (caffeine). The retrieved electrical impedance and video data were processed using Matlab. \ud \ud Main Outcome Measure(s):\ud The sperm beat frequency and amplitude determined from the electrical impedance and video data. \ud \ud Result(s):\ud The electrically measured sperm beat frequency was verified by optical analysis and in correspondence. Furthermore the microfluidic platform allowed single sperm analysis by altering the sperm by temperature and chemical stimuli. \ud \ud Conclusion(s):\ud This platform could be exploited as a potential tool to study sperm cells on the single cell level and to perform advanced sperm selection for intracytoplasmic sperm injection (ICSI) applications. (C) 2016 by American Society for Reproductive Medicine

Endothelial Dysfunction After ST-segment Elevation Myocardial Infarction and Long-term Outcome: A Study With Reactive Hyperemia Peripheral Artery Tonometry

Introduction and objectives: Long-term data on the relationship between endothelial dysfunction after ST-segment elevation myocardial infarction and future adverse clinical events are scarce. The aim of this study was to noninvasively assess whether endothelial dysfunction 4 to 6 weeks after primary percutaneous coronary intervention for acute ST-segment elevation myocardial infarction predicts future clinical events. Methods: This prospective cohort study was performed in 70 patients of the RESPONSE randomized trial, who underwent noninvasive assessment of endothelial function 4 to 6 weeks after primary percutaneous coronary intervention. Endothelial function was measured by the reactive hyperemia peripheral artery tonometry method; an index < 1.67 identified endothelial dysfunction. Results: The reactive hyperemia peripheral artery tonometry index measured on average 1.90 ± 0.58. A total of 35 (50%) patients had endothelial dysfunction and 35 (50%) patients had normal endothelial function. Periprocedural “complications” (eg, cardiogenic shock, total atrioventricular block) were more common in patients with endothelial dysfunction than in those without (25.7% vs 2.9%; P < .01). During 4.0 ± 1.7 years of follow-up, 20 (28.6%) patients had major adverse cardiovascular events: events occurred in 9 (25.7%) patients with endothelial dysfunction and in 11 (31.5%) patients with normal endothelial function (P = .52). There was an association between the prevalence of diabetes mellitus at baseline and the occurrence of major adverse cardiovascular events during follow-up (univariate analysis: hazard ratio = 2.8; 95% confidence interval, 1.0-7.8; P < .05), and even in multivariate analyses the risk appeared to be increased, although not significantly (multivariate analysis: hazard ratio = 2.5; 95% confidence interval, 0.8-7.5). Conclusions: In this series of patients who survived an ST-segment elevation myocardial infarction, endothelial dysfunction, as assessed by reactive hyperemia peripheral artery tonometry 4 to 6 weeks after myocardial infarction, did not predict future clinical events during a mean follow-up of 4 years

Endothelial function after ST-elevation myocardial infarction in patients with high levels of high-sensitivity CRP and Lp-PLA2: A substudy of the RESPONSE randomized trial

The combination of high levels of high-sensitive C-reactive protein (hs-CRP) and lipoprotein-associated phospholipase-A2 (Lp-PLA2) was recently shown to correlate with increased cardiovascular risk. Endothelial dysfunction is also known to be a risk factor for cardiovascular events. To test among patients with previous ST-elevation myocardial infarction (STEMI) the hypothesis that high levels of both hs-CRP and Lp-PLA2 may be associated with impaired endothelium-dependent vasodilatation. In this substudy of the RESPONSE randomized trial, we used reactive hyperemia peripheral artery tonometry (RH-PAT) 4 to 6weeks after STEMI and primary percutaneous coronary intervention (PPCI) to non-invasively assess endothelial function (RH-PAT index 239μg/L, respectively). Patients were 57.4±9.7years and 53 (77.9%) were men. 11 (16%) patients were classified as high-risk and 57 (84%) as low-to-intermediate-risk. The RH-PAT index was 1.68±0.22 in high-risk and 1.95±0.63 in low-to-intermediate-risk patients (p=0.17). Endothelial dysfunction was present in 8 (72.7%) high-risk and 26 (45.6%) low-to-intermediate-risk patients (p=0.09). Framingham risk score, NT-proBNP and fibrinogen levels were higher in high-risk patients (p≤0.03). In this population of patients with recent STEMI and PPCI, we observed between patients with high hs-CRP and Lp-PLA levels and all other patients no more than numerical differences in endothelial function that did not reach a statistical significance. Nevertheless, further research in larger study populations may be warrante

Endothelial Dysfunction After ST-segment Elevation Myocardial Infarction and Long-term Outcome: A Study With Reactive Hyperemia Peripheral Artery Tonometry

Long-term data on the relationship between endothelial dysfunction after ST-segment elevation myocardial infarction and future adverse clinical events are scarce. The aim of this study was to noninvasively assess whether endothelial dysfunction 4 weeks to 6 weeks after primary percutaneous coronary intervention for acute ST-segment elevation myocardial infarction predicts future clinical events. This prospective cohort study was performed in 70 patients of the RESPONSE randomized trial, who underwent noninvasive assessment of endothelial function 4 weeks to 6 weeks after primary percutaneous coronary intervention. Endothelial function was measured by the reactive hyperemia peripheral artery tonometry method; an index <1.67 identified endothelial dysfunction. The reactive hyperemia peripheral artery tonometry index measured on average 1.90±0.58. A total of 35 (50%) patients had endothelial dysfunction and 35 (50%) patients had normal endothelial function. Periprocedural "complications" (eg, cardiogenic shock, total atrioventricular block) were more common in patients with endothelial dysfunction than in those without (25.7% vs 2.9%; P <.01). During 4.0±1.7 years of follow-up, 20 (28.6%) patients had major adverse cardiovascular events: events occurred in 9 (25.7%) patients with endothelial dysfunction and in 11 (31.5%) patients with normal endothelial function (P=.52). There was an association between the prevalence of diabetes mellitus at baseline and the occurrence of major adverse cardiovascular events during follow-up (univariate analysis: hazard ratio=2.8; 95% confidence interval, 1.0-7.8; P <.05), and even in multivariate analyses the risk appeared to be increased, although not significantly (multivariate analysis: hazard ratio=2.5; 95% confidence interval, 0.8-7.5). In this series of patients who survived an ST-segment elevation myocardial infarction, endothelial dysfunction, as assessed by reactive hyperemia peripheral artery tonometry 4 weeks to 6 weeks after myocardial infarction, did not predict future clinical events during a mean follow-up of 4 year