Semantic segmentation of human oocyte images using deep neural networks.

BACKGROUND: Infertility is a significant problem of humanity. In vitro fertilisation is one of the most effective and frequently applied ART methods. The effectiveness IVF depends on the assessment and selection of gametes and embryo with the highest developmental potential. The subjective nature of morphological assessment of oocytes and embryos is still one of the main reasons for seeking effective and objective methods for assessing quality in automatic manner. The most promising methods to automatic classification of oocytes and embryos are based on image analysis aided by machine learning techniques. The special attention is paid on deep neural networks that can be used as classifiers solving the problem of automatic assessment of the oocytes/embryos. METHODS: This paper deals with semantic segmentation of human oocyte images using deep neural networks in order to develop new version of the predefined neural networks. Deep semantic oocyte segmentation networks can be seen as medically oriented predefined networks understanding the content of the image. The research presented in the paper is focused on the performance comparison of different types of convolutional neural networks for semantic oocyte segmentation. In the case study, the merits and limitations of the selected deep neural networks are analysed. RESULTS: 71 deep neural models were analysed. The best score was obtained for one of the variants of DeepLab-v3-ResNet-18 model, when the training accuracy (Acc) reached about 85% for training patterns and 79% for validation ones. The weighted intersection over union (wIoU) and global accuracy (gAcc) for test patterns were calculated, as well. The obtained values of these quality measures were 0,897 and 0.93, respectively. CONCLUSION: The obtained results prove that the proposed approach can be applied to create deep neural models for semantic oocyte segmentation with the high accuracy guaranteeing their usage as the predefined networks in other tasks.

Longitudinal study on novel neuropeptides phoenixin, spexin and kisspeptin in adolescent inpatients with anorexia nervosa - association with psychiatric symptoms.

OBJECTIVES: It is hypothesized that novel neuropeptides such as phoenixin (PNX), spexin (SPX), and kisspeptin (KISS) are involved in the pathogenesis of eating disorders. The study presented here analyzed neuropeptide concentrations during the course of anorexia nervosa (AN) and aimed to correlate those values with anthropometric and psychometric measurements. METHODS: A longitudinal study was carried outin 30 AN adolescent patients and 15 age-matched healthy female controls. Selected neuroprotein serum levels were analyzed in malnourished patients (accAN) and following partial weight recovery (norAN), and these values were compared with the control group. RESULTS: In accAN patients, decreased serum PNX levels were detected while SPX serum concentrations were lower in the accAN and norAN patients. No differences were observed in KISS concentrations in all studied groups. CONCLUSIONS: In malnourished adolescent inpatients with AN, serum PNX and SPX level were decreased. The partial weight recovery normalized PNX concentrations but failed to normalize SPX levels. Therefore these two neuropeptides might be crucial for the etiology and course of the AN. The KISS levels did not change in the course of AN. The PNX levels were associated with some symptoms of eating disorders which may indicate its potential contribution in the regulation of emotions and behaviors in AN.

Escitalopram as a modulator of proopiomelanocortin, kisspeptin, Kiss1R and MCHR1 gene expressions in the male rat brain.

Neuropeptides are important, multifunctional regulatory factors of the nervous system, being considered as a novel, atypical sites of antidepressants action. It has already been proven that some of them, such as selective serotonin reuptake inhibitors (SSRI), are able to affect peptidergic pathways in various brain regions. Despite these reports, there is so far no reports regarding the effect of treatment with SSRIs on brain proopiomelanocortin (POMC), kisspeptin, Kiss1R and MCHR1 gene expression. In the current study we examined POMC, kisspeptin, Kiss1R and MCHR1 mRNA expression in the selected brain structures (hypothalamus, hippocampus, amygdala, striatum, cerebellum and brainstem) of rats chronically treated with a 10 mg/kg dose of escitalopram using quantitative Real-Time PCR. Long-term treatment with escitalopram led to the upregulation of MCHR1 expression in the rat amygdala. Kisspeptin mRNA level was also increased in the amygdala, but Kiss1R mRNA expressions were elevated in the hippocampus, hypothalamus and cerebellum. POMC mRNA expressions were in turn decreased in the hippocampus, amygdala, cerebellum and brainstem. These results may support the hypothesis that these neuropeptides may be involved in the site-dependent actions of SSRI antidepressants. This is the first report of the effects of escitalopram on POMC, kisspeptin, Kiss1R and MCHR1 in animal brain. Our findings shed a new light on the pharmacology of SSRIs and may contribute to a better understanding of the alternative, neuropeptide-dependent modes of antidepressant action.

The first identification of nesfatin-1-expressing neurons in the human bed nucleus of the stria terminalis.

Neuropeptides are involved in various brain activities being able to control a wide spectrum of higher mental functions. The purpose of this concise structural investigation was to detect the possible immunoreactivity of the novel multifunctional neuropeptide nesfatin-1 within the human bed nucleus of the stria terminalis (BNST). The BNST is involved in the mechanism of fear learning, integration of stress and reward circuits, and pathogenesis of addiction. Nesfatin-1-expressing neurons were identified for the first time in several regions of the BNST using both immunohistochemical and fluorescent methods. This may implicate a potential contribution of this neuropeptide to the BNST-related mechanisms of stress/reward responses in the human brain.

The GnRH analogues affect novel neuropeptide SMIM20/phoenixin and GPR173 receptor expressions in the female rat hypothalamic-pituitary-gonadal (HPG) axis.

The recently discovered peptide phoenixin (PNX) and its receptor GPR173 are novel factors that exhibit a large spectrum of regulatory activity, especially when considered as a central modulator of GnRH-related hormonal control of reproductive processes. It has been already proven that GnRH agonists and antagonists can modulate peptidergic signalling in the HPG axis. Despite these findings, there is so far no information regarding the influence of treatment with GnRH analogues on SMIM20/phoenixin signalling in the hypothalamic-pituitary-gonadal axis. In the current study, SMIM20/phoenixin and GPR173 mRNA levels were measured in the hypothalamus, pituitary and ovaries of female rats in the dioestrus phase following treatment with GnRH-R agonist (buserelin) and antagonist (cetrorelix) using quantitative real-time PCR. The serum PNX concentrations were also estimated with ELISA technique. The hypothalamic, hypophyseal and especially ovarian levels of SMIM20 mRNA were increased after both buserelin and cetrorelix administration. The GPR173 expressions were in turn decreased in the hypothalamus and pituitary. Treatment with the GnRH analogues led to the modulation of SMIM20/phoenixin and GPR173 mRNA expression in the female rat hypothalamic-pituitary-gonadal axis. By identifying buserelin and cetrorelix as novel modulators of phoenixin signalling in the animal HPG axis, these results cast new light on the GnRH analogues mode of action and contribute to a better understanding of the mechanisms responsible for the hormonal control of reproduction.

Effect of long-term treatment with classical neuroleptics on NPQ/spexin, kisspeptin and POMC mRNA expression in the male rat amygdala.

Neuroleptics modulate the expression level of some regulatory neuropeptides in the brain. However, if these therapeutics influence the peptidergic circuits in the amygdala remains unclear. This study specifies the impact profile of the classical antipsychotic drugs on mRNA expression of the spexin/NPQ, kisspeptin-1 and POMC in the rat amygdala. Animals were treated with haloperidol and chlorpromazine for 28 days prior to transcript quantification via qPCR. Haloperidol and chlorpromazine induced a change in the expression of all neuropeptides analyzed. Both drugs led to the decrease of Kiss-1 expression, whereas in POMC and spexin/NPQ their up-regulation in the amygdala was detected. These modulating effects on may represent alternative, so far unknown mechanisms, of classical antipsychotic drugs triggering pharmacological responses.

Neuroleptics Affect Kisspeptin mRNA Expression in the Male Rat Hypothalamus and Hippocampus.

Introduction: Kisspeptin has a multidirectional neuroendocrinal activity. It is especially considered to be a central regulator of reproductive function. Numerous data proved that neuroleptic administration may affect the peptidergic signaling in the various brain structures. However, there is no information concerning the relationship between treatment with neuroleptics and brain kisspeptin mRNA expression. Methods: We assessed the kisspeptin mRNA level in the hypothalamus and hippocampus of rats shortly and chronically (28 days) treated with haloperidol, chlorpromazine, and olanzapine using a quantitative Real-Time PCR method. Results: We have shown that all studied neuroleptics injected chronically have the ability to downregulate the kisspeptin mRNA expression in the hypothalamus, which may suggest the presence of an alternative mechanism for their orexigenic side effects. Long-term treatment with chlorpromazine increased the level of kisspeptin mRNA expression in the hippocampus. Discussion: Our results shed a new light on the pharmacology of antipsychotics and may contribute to a better understanding of alternative mechanisms responsible for their action. The study also highlights a complex nature of potential connections between dopamine transmission and brain kisspeptin pathways.

Long-term treatment with haloperidol affects neuropeptide S and NPSR mRNA levels in the rat brain.

OBJECTIVE: The brainstem-derived neuropeptide S (NPS) has a multidirectional regulatory activity, especially as a potent anxiolytic factor. Accumulating data suggests that neuroleptics affect peptidergic signalling in various brain structures. However, there is no information regarding the influence of haloperidol on NPS and NPS receptor (NPSR) expression. METHODS: We assessed NPS and NPSR mRNA levels in brains of rats treated with haloperidol using quantitative real-time polymerase chain reaction. RESULTS: Chronic haloperidol treatment (4 weeks) led to a striking upregulation of NPS and NPSR expression in the rat brainstem. Conversely, the NPSR mRNA expression was decreased in the hippocampus and striatum. CONCLUSIONS: This stark increase of NPS in response to haloperidol treatment supports the hypothesis that this neuropeptide is involved in the dopamine-dependent anxiolytic actions of neuroleptics and possibly also in the pathophysiology of mental disorders. Furthermore, our findings underline the complex nature of potential interactions between dopamine receptors and brain peptidergic pathways, which has potential clinical applications.

Neurolight -astonishing advances in brain imaging.

In recent years, significant advances in basic neuroanatomical studies have taken place. Moreover, such classical, clinically-oriented human brain imaging methods such as MRI, PET and DTI have been applied to small laboratory animals allowing improvement in current experimental neuroscience. Contemporary structural neurobiology also uses various technologies based on fluorescent proteins. One of these is optogenetics, which integrates physics, genetics and bioengineering to enable temporal precise control of electrical activity of specific neurons. Another important challenge in the field is the accurate imaging of complicated neural networks. To address this problem, three-dimensional reconstruction techniques and retrograde labeling with modified viruses has been developed. However, a revolutionary step was the invention of the "Brainbow" system, utilizing gene constructs including the sequences of fluorescent proteins and the usage of Cre recombinase to create dozens of colour combinations, enabling visualization of neurons and their connections in extremely high resolution. Furthermore, the newly- introduced CLARITY method should make it possible to visualize three-dimensionally the structure of translucent brain tissue using the hydrogel polymeric network. This original technique is a big advance in neuroscience creating novel viewpoints completely different than standard glass slide immunostaining.

Dual orexin receptor antagonists - promising agents in the treatment of sleep disorders.

Insomnia is a serious medical and social problem, its prevalence in the general population ranges from 9 to 35% depending on the country and assessment method. Often, patients are subject to inappropriate and therefore dangerous pharmacotherapies that include prolonged administration of hypnotic drugs, benzodiazepines and other GABAA receptor modulators. This usually does not lead to a satisfactory improvement in patients' clinical states and may cause lifelong drug dependence. Brain state transitions require the coordinated activity of numerous neuronal pathways and brain structures. It is thought that orexin-expressing neurons play a crucial role in this process. Due to their interaction with the sleep-wake-regulating neuronal population, they can activate vigilance-promoting regions and prevent unwanted sleep intrusions. Understanding the multiple orexin modulatory effects is crucial in the context of pathogenesis of insomnia and should lead to the development of novel treatments. An important step in this process was the synthesis of dual antagonists of orexin receptors. Crucially, these drugs, as opposed to benzodiazepines, do not change the sleep architecture and have limited side-effects. This new pharmacological approach might be the most appropriate to treat insomnia.

Hypothalamic subependymal niche: a novel site of the adult neurogenesis.

The discovery of undifferentiated, actively proliferating neural stem cells (NSCs) in the mature brain opened a brand new chapter in the contemporary neuroscience. Adult neurogenesis appears to occur in specific brain regions (including hypothalamus) throughout vertebrates' life, being considered an important player in the processes of memory, learning, and neural plasticity. In the adult mammalian brain, NSCs are located mainly in the subgranular zone (SGZ) of the hippocampal dentate gyrus and in the subventricular zone (SVZ) of the lateral ventricle ependymal wall. Besides these classical regions, hypothalamic neurogenesis occurring mainly along and beneath the third ventricle wall seems to be especially well documented. Neurogenic zones in SGZ, SVZ, and in the hypothalamus share some particular common features like similar cellular cytoarchitecture, vascularization pattern, and extracellular matrix properties. Hypothalamic neurogenic niche is formed mainly by four special types of radial glia-like tanycytes. They are characterized by distinct expression of some neural progenitor and stem cell markers. Moreover, there are numerous suggestions that newborn hypothalamic neurons have a significant ability to integrate into the local neural pathways and to play important physiological roles, especially in the energy balance regulation. Newly formed neurons in the hypothalamus can synthesize and release food intake regulating neuropeptides and they are sensitive to the leptin. On the other hand, high-fat diet positively influences hypothalamic neurogenesis in rodents. The nature of this intriguing new site of adult neurogenesis is still so far poorly studied and requires further investigations.

AMP-activated protein kinase is involved in induction of protective autophagy in astrocytes exposed to oxygen-glucose deprivation.

AMP-activated kinase (AMPK) acts as the intracellular ATP depletion sensor, which detects and limits increases in the AMP/ATP ratio. AMPK may be significantly activated under stress conditions that deplete cellular ATP levels such as ischemia/hypoxia or glucose deprivation. Recent studies strongly suggest that AMPK participates in autophagy regulation, but it is not known whether AMPK activated by ischemia regulates autophagy in astrocytes and the consequence of autophagy activation in ischemic astrocytes are unclear. We have investigated the contribution of AMPK to autophagy activation in rat primary astrocyte cultures subjected to ischemia-simulating conditions (combined oxygen glucose deprivation, OGD) and its potential effects on astrocyte damage induced by OGD (1-12 h). The evidence supports the conclusion that AMPK activation at early stages of OGD is involved in induction of protective autophagy in astrocytes. Inhibition of AMPK, either by siAMPKα1 or by compound C, significantly attenuated the expression of autophagy-related proteins and decrease of astrocyte viability following OGD. The findings provide additional data about the role of AMPK in ischemic astrocytes and downstream responses that may be involved in OGD-induced protective autophagy.

NiTi Shape Memory Clamps with Modified Surface for Bone Fracture Treatment.

Using NiTi alloys with shape memory for long-term medical implants requires modification of their surface due to the possible occurrence of corrosion. Hence, the surface of the staples used to join fractured bone within the craniofacial region was modified by applying a titanium oxy-nitrogen layer and a hydroxyapatite coating. Surface-modified clamps were tested in vivo using New Zealand white rabbits. After determining the mechanical characteristics of the bone and considering the initial state and surface modification, the diameter of the wire (used to make the clamps with the appropriate compression force) was selected. Implantation was performed on two groups of rabbits: experimental and control. In the experimental group, an intentionally induced bone fracture was treated in one tibia. On the second tibia, two additional clamps were applied to increase the possibility of a negative impact of the NiTi alloy on a living organism. After 6 weeks of application, a proper joining of the broken bone fragments was stated. Whereas after twelve weeks, no negative impact of the clamp material on a living organism, i.e., a rabbit, was found. Hence, the clamp with the modified surface can connect bone fragments in humans as well as small and medium-sized animals, with an extended range of use up to 12 weeks.

Morphological and enzymatic changes caused by a long-term treatment of female rats with a low dose of gonadoliberin agonist and antagonist.

BACKGROUND: Long-term treatment with gonadoliberin analogs is used to block the hypothalamic-pituitary-gonadal axis. The use of these agents is generally considered to be safe; however, some observations suggest the possibility of adverse effects. MATERIAL/METHODS: We investigated whether a 3-months administration of a low dose (6 µg/kg b.w.) of dalarelin - a new agonist, and cetrorelix - a known antagonist of GnRH to female rats causes morphological changes in pituitary gland, ovaries, uterus and liver (HE and VG staining); effects on pituitary, hepatic and blood enzyme activities (histochemical and kinetic methods, respectively), and on the blood lipid profile (colorimetric methods); and to what extent these changes are reversible. RESULTS: Applying analogs effectively inhibited ovulation, affected the uterine endometrium and changed histological appearance of the liver (e.g., steatosis). They altered activities of marker enzymes of cellular respiration, gluconeogenesis and intracellular digestion in the liver and, partially in the pituitary gland, caused undesirable changes in the activities of aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, and creatine kinase, and a concentration of cholesterol HDL fraction and triglycerides in the blood. Both morphological and enzymatic effects were more evident after antagonist administration; changes in the blood lipid profile were more evident after agonist administration. In both analogs histological and enzymatic changes persisted a relatively long time after the discontinuation of the treatment. CONCLUSIONS: The low dose of dalarelin and cetrorelix is sufficient to cause limited damage of hepatic cells and may modify the function of pituitary, ovaries, uterus and liver as well as other organs, even after discontinuation of the treatment.

Profiles of interferon-gamma and interleukin-2 in patients after allogeneic hematopoietic stem cell transplantation.

BACKGROUND: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) may be related to the occurrence of complications, including graft-versus-host disease (GvHD) and infections. The pathogenesis of acute GvHD is connected with T lymphocytes, which identify alloantigens on host's antigen-presenting cells, activate production of interferon-gamma (IFN-gamma) and interleukin-2 (IL-2), and act on the immune effector cells and damage tissues and organs. AIM: The aim of the study was to investigate and distinguish serum concentration profiles of IFN-gamma and IL-2 within a 30-d period after allo-HSCT. METHODS: We enrolled 62 patients, i.e., 30 (48%) male and 32 (52%) female subjects [median age 49.5 (19-68) years], after allo-HSCT from siblings (n = 12) or unrelated donors (n = 50) due to acute myeloid leukemia with myeloablative conditioning (n = 26; 42%) and with non-myeloablative conditioning (n = 36; 58%). All patients were given standard immunosuppressive therapy with cyclosporin-A and methotrexate and pre-transplant antithymocyte globulin in the unrelated setting. Blood samples were collected pre-transplant before and after (on day -1) the conditioning therapy and on days +2,+4, +6, +10, +20, and +30 after allo-HSCT. Serum levels of IL-2 and IFN-gamma were determined using ELISA. RESULTS: Patients were divided into four groups depending on the presence of acute GvHD and clinical manifestations of infection. Group I included patients with neither acute GvHD nor infections [n = 15 (24%)], group II consisted of patients with infections without acute GvHD [n = 17 (27%)], group III was comprised of patients with acute GvHD without infections [n = 9 (15%)], and group IV included patients with both acute GvHD and infections [n = 21 (34%)]. IFN-gamma concentrations were higher in Group II than in other groups on days +20 (P = 0.014) and +30 (P = 0.008). Post-hoc tests showed lower concentrations of IFN-gamma on day +30 in groups I (P = 0.039) and IV (P = 0.017) compared to group II. The levels of IL-2 were mostly undetectable. CONCLUSION: Serum levels of IFN-gamma following allo-HSCT progressively escalate. High serum levels of IFN-gamma are related to infectious complications rather than acute GvHD. Serum concentrations of IL-2 in most patients are undetectable.

Post-tracheostomy complications: respiratory failure caused by authologic foreign body-case report.

Tracheostomy is performed frequently as a palliative treatment in patients with end-stage respiratory failure (RF). However, in patients requiring prolonged mechanical ventilation it may be difficult to recognize and can often lead to life-threatening RF. We present two cases of acute-on-chronic respiratory failure (ACRF) occurring in patients who had undergone tracheostomy [one with percutaneous dilatational tracheostomy (PDT) and the second with surgical tracheostomy (ST)]. The first case was admitted due to ACRF several months after previous successful decannulation and the second case after failure of several attempts of weaning from tracheal cannula. In both cases, noninvasive mechanical ventilation assisted flexible bronchoscopy (NIV-FB) was able to identify and solve the tracheal stenosis secondary to stiff banana-shaped whitish foreign bodies. Histology sampling and genetic testing confirmed autologous foreign body formation-tracheal cartilage calcification. NIV-FB was found to be safe and effective in both diagnosis and treatment of the tracheal stenosis. Life-threatening RF connected with tracheal stenosis may be caused by rupture of tracheal cartilage ossification in patients with a history of ST and PDT. Bronchofiberoscopy performed with NIV will be a useful procedure to evaluate and treat the respiratory tract in patients with RF with suspected tracheal stenosis.

Eotaxin, but not IL-8, is increased in upper and lower airways of allergic rhinitis subjects after nasal allergen challenge.

The aim of this study was to assess the impact of a single nasal allergen challenge (NAC) on levels of eotaxin and IL-8 and the inflammatory cells in upper and lower airways of allergic rhinitis (AR) patients. Twenty-four AR patients and 12 control subjects entered a sequential nasal placebo challenge and NAC study, out of the pollen season. Nasal lavage fluid (NLF) was obtained at baseline, 15 minutes, and 1, 5, and 24 hours postchallenge. Before and 24 hours after placebo/allergen challenge induced sputum was performed. NLF and induced sputum were evaluated for total cell count (TCC) and differential cell count and analyzed for concentrations of eotaxin and IL-8 using ELISA method. NAC in AR subjects was associated with significantly increased sputum (p = 0.008) and NLF (p < 0.001) eotaxin levels. Post-NAC IL-8 levels were significantly increased in NLF (p < 00001) but not in sputum (p = 0.080) of AR subjects. Increased eotaxin levels in NLF positively correlated with the increased TCC and eosinophils. Positive correlations were also found between NLF increased eotaxin level and sputum TCC, eosinophils, and macrophages. NAC is associated with the increased levels of eotaxin in lower airways of AR subjects. Allergen-induced secretion of eotaxin in nasal mucosa of AR subjects is involved in determining the cellular character of both upper and lower airway inflammation.

[Autophagy in brain ischemia]. / Autofagia w niedokrwieniu mózgu.

Autophagy is an intracellular process of macromolecule and organelle degradation, which plays an important role both in maintaining homeostasis and in responding to various harmful stimuli. Recent studies clearly indicate upregulation of autophagy in neurons challenged with brain ischemia. In this paper we present biosynthesis of autophagosomes as well as the role and molecular mechanisms of basal and induced neuronal autophagy. We have also reviewed recently published papers concerning the potential role of autophagy in brain ischemia. Results of both in vivo and in vitro experimental studies indicate that signaling pathways related to autophagy might become a target of new neuroprotective strategies.

[S-adenosyl L-methionine in CNS diseases]. / S-adenozylo L-metionina w schorzeniach OUN.

S-adenosyl L-methionine (SAMe) is the natural, universal methyl group donor, participating in transmethylation reactions, known and commonly used as a dietary supplement since 1952. It plays an important role in the synthesis of neuromediators and melatonin and mechanisms of epigenetic regulation. The aim of this article is to review the literature about possibilities of SAMe application in the therapy of CNS diseases: depression, dementia syndromes, schizophrenia and somatic disorders. SAMe is the promising dietary supplement, which may be successfully used as a substance increasing effectiveness of the treatment of depression, with antidepressants in monotherapy in mild depressive states or depressive symptoms. SAMe addition to antipsychotic drug, may lead to the improvement of the quality of life and reduction of aggressiveness of patients. SAMe may be an effective substance in the therapy and prophylaxis of mild cognitive impairments and mild dementia syndrome. SAMe possesses some hepatoprotective action, so it may decrease the risk of the development of neoplasm, alcohol-induced liver disease (ALD) and cirrhosis. SAMe improves the functions of joints and decreases the experience of pain in rheumatoid arthritis (RA).