Endocytosis and Alzheimer's disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder and is the most common cause of dementia. The pathogenesis of AD still remains unclear, including two main hypotheses: amyloid cascade and tau hyperphosphorylation. The hallmark neuropathological changes of AD are extracellular deposits of amyloid-ß (Aß) plaques and intracellular neurofibrillary tangles (NFTs). Endocytosis plays an important role in a number of cellular processes including communication with the extracellular environment, nutrient uptake, and signaling by the cell surface receptors. Based on the results of genetic and biochemical studies, there is a link between neuronal endosomal function and AD pathology. Taking this into account, we can state that in the results of previous research, endolysosomal abnormality is an important cause of neuronal lesions in the brain. Endocytosis is a central pathway involved in the regulation of the degradation of amyloidogenic components. The results of the studies suggest that a correlation between alteration in the endocytosis process and associated protein expression progresses AD. In this article, we discuss the current knowledge about endosomal abnormalities in AD.

Small extracellular vesicles - A host for advanced bioengineering and "Trojan Horse" of non-coding RNAs.

Small extracellular vesicles (sEVs) are a type of membranous vesicles that can be released by cells into the extracellular space. The relationship between sEVs and non-coding RNAs (ncRNAs) is highly intricate and interdependent. This symbiotic relationship plays a pivotal role in facilitating intercellular communication and holds profound implications for a myriad of biological processes. The concept of sEVs and their ncRNA cargo as a "Trojan Horse" highlights their remarkable capacity to traverse biological barriers and surreptitiously deliver their cargo to target cells, evading detection by the host-immune system. Accumulating evidence suggests that sEVs may be harnessed as carriers to ferry therapeutic ncRNAs capable of selectively silencing disease-driving genes, particularly in conditions such as cancer. This approach presents several advantages over conventional drug delivery methods, opening up new possibilities for targeted therapy and improved treatment outcomes. However, the utilization of sEVs and ncRNAs as therapeutic agents raises valid concerns regarding the possibility of unforeseen consequences and unintended impacts that may emerge from their application. It is important to consider the fundamental attributes of sEVs and ncRNAs, including by an in-depth analysis of the practical and clinical potentials of exosomes, serving as a representative model for sEVs encapsulating ncRNAs.

Quantitative Phase Imaging Detecting the Hypoxia-Induced Patterns in Healthy and Neoplastic Human Colonic Epithelial Cells.

Hypoxia is a frequent phenomenon during carcinogenesis and may lead to functional and structural changes in proliferating cancer cells. Colorectal cancer (CRC) is one of the most common neoplasms in which hypoxia is associated with progression. The aim of this study was to assess the optical parameters and microanatomy of CRC and the normal intestinal epithelium cells using the digital holotomography (DHT) method. The examination was conducted on cancer (HT-29, LoVo) and normal colonic cells (CCD-18Co) cultured in normoxic and hypoxic environments. The assessment included optical parameters such as the refractive index (RI) and dry mass as well as the morphological features. Hypoxia decreased the RI in all cells as well as in their cytoplasm, nucleus, and nucleoli. The opposite tendency was noted for spheroid-vesicular structures, where the RI was higher for the hypoxic state. The total volume of hypoxic CCD-18Co and LoVo cells was decreased, while an increase in this parameter was observed for HT-29 cells. Hypoxia increased the radius and cell volume, including the dry mass of the vesicular content. The changes in the optics and morphology of hypoxic cells may suggest the possibility of using DHT in the detection of circulating tumor cells (CTCs).

Label-Free Quantitative Phase Imaging Reveals Spatial Heterogeneity of Extracellular Vesicles in Select Colon Disorders.

Three-dimensional (3D) imaging and quantitative analysis of extracellular vesicles (EVs) remain largely unexplored, mainly because of limitations in detection techniques. In this study, EVs from patients diagnosed with colorectal cancer (CRC) and ulcerative colitis were examined. To investigate the spatial heterogeneity and 3D refractive index (RI) distribution of single EVs, a label-free digital holographic tomography technique was used at a submicrometer spatial resolution. The presented image-processing algorithms were used in quantitative analysis with digital staining and 3D visualization, the determination of the EV size distribution and extraction of fractions with different RIs. Reconstructed 3D RI distributions revealed variations in the spatial heterogeneity of EVs related to tissue specificity, such as CRC, normal colonic mucosa, and ulcerative colitis, as well as the isolation procedures used. The RI values of EVs isolated from solid tissues of frozen CRC samples were also dependent on the tumor grade and cancer cell proliferation. The simultaneous examination of cell culture models confirmed the association of the RI of EVs with the tumor grade. 3D-RI data analysis generates new perspectives with the optical, contact-free, label-free examination of the individual EVs. Depending on the specific tissue and isolation method, EVs exhibit significant spatial heterogeneity. The optical parameters of single EVs enabled their classification into two unique subgroups with different RI values.

Molecular profiling of the intestinal mucosa and immune cells of the colon by multi-parametric histological techniques.

The impact of the post-mortem interval (PMI) on the optical molecular characteristics of the colonic mucosa and the gut-associated lymphoid tissue (GALT) were examined by multi-parametric measurements techniques. Inflammatory cells were identified by immunohistochemical staining. Molecular parameters were estimated using the Raman spectroscopy (RS) and Fourier Transform Infrared (FTIR) spectroscopic imaging. The 3D refractive index (3D-RI) distributions of samples were determined using the digital holographic tomography. The distribution of immune cells between post-mortem (PM) and normal controls did show significant differences for CD4 (P = 0.0016) or CD8 (P < 0.0001), whose expression level was decreased in PM cases. No association was found between individual PMI values and inflammatory cell distribution. However, there was a tendency for a negative correlation between CD4+ cells and PMI (r = - 0.542, P = 0.032). The alterations ongoing in post-mortem tissue may suggest that PMI has a suppressive effect on the effector properties of the cell-mediated immunity. Moreover, it was confirmed that spectroscopic and digital holotomographic histology are also a useful technique for characterization of the differences in inflammation of varying intensity and in GALT imaging in a solid tissue. Anatomical location of immune cells and methods of tissue fixation determine the molecular and optical parameters of the examined cases.

Interplay of stromal tumor-infiltrating lymphocytes, normal colonic mucosa, cancer-associated fibroblasts, clinicopathological data and the immunoregulatory molecules of patients diagnosed with colorectal cancer.

A total of 94 patients with colorectal cancer (CRC) were included in this study. Lymphocytic infiltration of CD45+ cells in the normal colon was more pronounced than that in the paired tumor stroma (p = 0.0008). The mean immunoscore of CD45+TILs was decreased in CRC compared with the controls (p = 0.0010). The percentage of CD3+ cells was higher in stage II than in stage IV (p = 0.0218) and showed a negative correlation with the TNM classification (r = -0.2867, p = 0.0109). The number of stromal CD4+TILs was higher in stage I than in stage III (p = 0.0116) and IV (p = 0.0104), and there was a negative correlation between this number and the stage (r = -0.3708, p = 0.0008). There was a positive correlation between the Ki-67 and CD45+ (r = 0.2468, p = 0.0294), CD3+ (r = 0.3822, p = 0.0006), and CD4+ cells (r = 0.5465, p < 0.0001). The levels of cancer-associated fibroblast (CAF) markers such as α-SMA, thrombin and fibronectin were significantly higher in CRC than in normal colonic mucosa. The immunohistochemical expression of α-SMA was negatively correlated with TILs, while fibronectin showed positive coexpression. A higher number of cells expressing IL-2Rα, PD-L1, CD33 and CD14 were found in colorectal adenocarcinomas than in controls. The number of CD14+ cells was also dependent on the TNM stage (p = 0.0444) and tumor budding (p = 0.0324). These findings suggest a suppressive impact of CRC on the adaptive immune response and emphasize the importance of CAFs in regulating tumor immunity.

Comparative analysis of exosome markers and extracellular vesicles between colorectal cancer and cancer-associated normal colonic mucosa.

INTRODUCTION: Exosomes are currently considered as the new biomarkers of colorectal cancer (CRC). Tetraspanins (CD9, CD63) belong to the well­known exosome markers, but can also be found on other subtypes of extracellular vesicles (EVs). OBJECTIVES: The aim of this study was to estimate the expression level of exosome markers and EVs in CRC. PATIENTS AND METHODS: The expression level of CD9 and CD63 antigens was evaluated by immunohistochemical staining in 109 patients diagnosed with CRC. Immunohistochemistry results were verified by nanoparticle tracking analysis (NTA), as well as the Western blot analysis and transmission electron microscopy. Exosomes isolation was performed on solid tissues. The immunohistochemical expression of both tetraspanins was compared with expression of cellular p roliferation marker, Ki­67. RESULTS: A higher expression level of exosome markers was observed in CRC compared with the normal colonic mucosa. The NTA revealed higher concentrations of nanoparticles in CRC tissues than in controls. There was a strong positive correlation between exosome markers and the Ki­67 antigen. The expression levels of both tetraspanins were different for lymph node stagi ng (N stage). CONCLUSIONS: Exosome markers and EVs were more pronounced in the CRC samples compared with controls. Immunohistochemical evaluation of tetraspanins reflects the results obtained by the NTA. Exocytosis appears to play an important role in the pathogenesis of CRC. To the best of our knowledge, such analysis was carried out for the first time.

Modeling of the immune response in the pathogenesis of solid tumors and its prognostic significance.

BACKGROUND: Tumor initiation and subsequent progression are usually long-term processes, spread over time and conditioned by diverse aspects. Many cancers develop on the basis of chronic inflammation; however, despite dozens of years of research, little is known about the factors triggering neoplastic transformation under these conditions. Molecular characterization of both pathogenetic states, i.e., similarities and differences between chronic inflammation and cancer, is also poorly defined. The secretory activity of tumor cells may change the immunophenotype of immune cells and modify the extracellular microenvironment, which allows the bypass of host defense mechanisms and seems to have diagnostic and prognostic value. The phenomenon of immunosuppression is also present during chronic inflammation, and the development of cancer, due to its duration, predisposes patients to the promotion of chronic inflammation. The aim of our work was to discuss the above issues based on the latest scientific insights. A theoretical mechanism of cancer immunosuppression is also proposed. CONCLUSIONS: Development of solid tumors may occur both during acute and chronic phases of inflammation. Differences in the regulation of immune responses between precancerous states and the cancers resulting from them emphasize the importance of immunosuppressive factors in oncogenesis. Cancer cells may, through their secretory activity and extracellular transport mechanisms, enhance deterioration of the immune system which, in turn, may have prognostic implications.

Brain tumour homogenates analysed by surface-enhanced Raman spectroscopy: Discrimination among healthy and cancer cells.

One of the biggest challenge for modern medicine is to make a discrimination among healthy and cancerous tissues. Therefore, nowadays big effort of scientist are devoted to find a new way for as fast as possible diagnosis with as much as possible accuracy in distinguishing healthy from cancerous tissues. That issues are probably the most important in the case of brain tumours, when the diagnosis time plays a great role. Herein we present the surface-enhanced Raman spectroscopy (SERS) together with the principal component analysis (PCA) used to identify the spectra of different brain specimens, healthy and tumour tissues homogenates. The presented analyses include three sets of brain tissues as control samples taken from healthy objects (one set consists of samples from four brain lobes and both hemispheres; eight samples) and the brain tumours from five patients (two Anaplastic Astrocytoma and three Glioblastoma samples). Results prove that tumour brain samples can be discriminated well from the healthy tissues by using only three main principal components, with 96% of accuracy. The largest influence onto the calculated separation is attributed to the spectral regions corresponding in SERS spectra to vibrations of the L-Tryptophan (1450, 1278 cm-1), protein (1300 cm-1), phenylalanine and Amide-I (1005, 1654 cm-1). Therefore, the presented method may open the way for the probable application as a very fast diagnosis tool alternative for conventionally used histopathology or even more as an intraoperative diagnostic tool during brain tumour surgery.

The expression of IL10RA in colorectal cancer and its correlation with the proliferation index and the clinical stage of the disease.

INTRODUCTION: Despite the widely described role of IL10 in immune response regulation during carcinogenesis, there is no established model describing the role of its receptor. The aim of this study is to elucidate the relationship between the subunit alpha of IL10 receptor (IL10RA) in the pathogenesis of colorectal cancer (CRC). METHODS: The study was conducted on archived paraffin blocks of 125 CRC patients, from which tissue microarrays (TMA) were made. These were subsequently used for immunohistochemistry to assess the expression of IL10RA, IL10, phosphorylated STAT3 (pSTAT3) and the Ki67 proliferation index. The intensity of both reactions was assessed by independent researchers using two approaches: digital image analysis and the Remmele and Stegner score (IRS). To assess the possible correlations between the two investigated markers and the clinical stage of CRC, the Pearson correlation coefficient was calculated. The expression of aforementioned proteins was assessed in tumor samples, healthy surgical margins and healthy control samples, obtained from cadavers during autopsy from the Department of Forensic Medicine. Statistical analysis was conducted using Statistica ver. 13.05 software. RESULTS: The final analysis included 105 CRC patients with complete clinical and pathological data, for whom the expressions of IL10RA, IL10, pSTAT3 and Ki67 were assessed using two independent methods. There was a positive correlation between the IL10RA expression and Ki-67 proliferation index (R = 0.63, p < 0.001) and a negative correlation between the IL10RA expression and the clinical stage of CRC (R = -0.21, p = 0.022). IL10RA correlated positively with pSTAT3 and IL10 in neoplastic tissue and tumor margin (with p < 0.01 for all correlations). We also observed a significantly higher expression of IL10RA in healthy surgical margins when compared to the actual tumor (p = 0.023, the paired t-test). The expression of IL10 was significantly higher in tumors than in healthy intestinal endothelium from control group. CONCLUSIONS: The correlations between the expression of IL10RA and the proliferation index or the clinical stage of CRC seem to confirm the importance of IL10RA in the pathogenesis of CRC. The higher expression of IL10RA in healthy surgical margins than in the tumor itself may suggest that IL10RA plays a role in regulating immune response to the neoplasm.

Association Between Interleukin-10 Receptors and the CD45-Immunophenotype of Central Nervous System Tumors: A Preliminary Study.

BACKGROUND/AIM: One of the current hypotheses assumes that brain tumors exert an immunosuppressive influence on the surrounding cellular environment. Interleukin-10 (IL-10) is one of the immunosuppressive cytokines modifying the biological activity of cancer. The aim of this study was to assess the expression of IL10R in CD45+ cells within primary brain tumors and metastases and establish its association with tumor basic immunophenotype. PATIENTS AND METHODS: Tissue samples were obtained intraoperatively during surgeries of 32 patients suffering from meningiomas (n=9), gliomas (n=12) and metastatic tumors (n=11). Expression was assessed with flow cytometry and immunohistochemical reactions. RESULTS: Expression of IL10R subunits within the leukocyte population (CD45+ cells) was significantly higher in primary tumors than in metastases. CONCLUSION: To the best of our knowledge, this is the first study describing a correlation between the IL10R expression on leukocytes and histological types of brain tumors.

Clinical Phenotype of Depression Affects Interleukin-6 Synthesis.

Major depressive disorder (MDD) is not a single disease, but a number of various ailments that form one entity. Psychomotor retardation, anhedonia, sleep disorders, an increased suicide risk, and anxiety are the main symptoms that often define the clinical diagnosis of depression. Interleukin-6 (IL-6), as one of the proinflammatory cytokines, seems to be overexpressed during certain mental disorders, including MDD. Overexpression of IL-6 in depression is thought to be a factor associated with bad prognosis and worse disease course. IL-6 may directly affect brain functioning and production of neurotransmitters; moreover, its concentration is correlated with certain clinical symptoms within the wide range of depressive symptomatology. Furthermore, there is a strong correlation between IL-6 synthesis and psychosomatic functioning of the patient. This article discusses potential sources and significance of IL-6 in the pathogenesis of depression.

LEVOMILNACIPRAN--A SUCCESSOR OF MILNACIPRAN WITH A HIGHER NORADRENERGIC SELECTIVITY.

A new antidepressant, levomilnacipran, is the levorotatory enantiomer of milnacipran. The drug belongs to selective serotonin-norepinephrine reuptake inhibitors (SNRI) and has the highest noradrenergic selectivity of all members of this group of antidepressants. Clinical trials have confirmed the effectiveness of levomilnacipran in the treatment of depression. The drug was placed on the US market in the form of prolonged-release capsules, which greatly simplifies the treatment of psychiatric patients. The safety of the drug is also higher than the safety of a racemate, resulting in a beneficial impact on the therapeutic effect. In this paper we present current information on the pharmacological and clinical properties of the newest antidepressant--levomilnacipran.

Compulsive buying in outline.

In spite of a hundred year long history of scientific research compulsive buying has been a hardly known phenomenon until today. Ambiguous scientific information makes it impossible to classify compulsive buying as a separate mental disorder. Recently many researchers have noticed phenomenological compatibility of compulsive buying with behavioural addictions. Nowadays, there is reasonable grounds that compulsive buying disorder can be defined as an addiction. There are many similarities occurring between a consumer type behaviours in compulsive buyers and a pathologic consumption of psychoactive substances which included the obsessive need to consumer or a compulsion to consume, personal dependence and loss of control over self-behaviour, as well as tendencies to the consumption increase. Compulsive buying disorder differs in its course from the compulsive behaviours. A strong compulsion to make a given activity, often impossible to restrain is associated with overwhelming but acceptable desire to purchase a specific item. Due to the latest information about the described phenomenon, it has been decided to present current knowledge of adequate classifications, epidemiology and therapy of compulsive buyers. In the article authors' own standpoint as regards pathogenesis and potential risk factors was described.